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A HANDBOOK OX ANTISEPTICS 



THE MACMILLAN COMPANY 

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MACMILLAN & CO., Limited 

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TORONTO 



A HANDBOOK 

ON 

ANTISEPTICS 

BY 

HENRY DRYSDALE DAKJN, D.Sc, F.I.C., F.R.S. 

AND 

EDWARD KELLOGG DUNHAM, M.D. 

EMERITUS PROFESSOR OF PATHOLOGY, UNIVERSITY AND 

EELLEVUE HOSPITAL MEDICAL COLLEGE 
MAJOR, MEDICAL OFFICERS RESERVE CORPS, U. S. ARMY 



THE MACMILLAN COMPANY 
1917 

All rights reserved 



lb* 



Copyright, 1917, 
By THE MACMILLAN COMPANY. 

Set up and electrotyped. Published October, 1917. 



OCT 18 1317 

J. S. dishing- Co. — Berwick & Smith Co. 
Norwood, Mass., U.S.A. 

©CI. A 47665*5 



LOUIS PASTEUR JOSEPH LISTER 

1822-1895 1827-1913 

An Extract from Pasteur's Address at the 
Celebration of his Seventieth Birthday 

You lastly, delegates of foreign nations, who have come 
from so far to give proof of jour sympathy with France — you 
bring me the deepest happiness which a man can experience 
who believes implicitly that science and peace will triumph 
over ignorance and war, that people will learn to agree to- 
gether, not for purposes of destruction but for improvement, 
and that the future will belong to those who shall have done 
the most for suffering humanity. 

I address myself to you, my dear Lister, and to all of you 
illustrious representatives of Science, of Medicine, and of 
Surgery. 

Young people, young people, confine yourselves to those 
methods, sure and powerful, of which we as yet know only 
the first secrets. And all, however noble your career, never 
permit yourselves to be overcome by scepticism*, both un- 
worthy and barren ; neither permit the hours of sadness which 
pass over a nation to discourage you. Live in the serene 
peace of your laboratories and your libraries. First ask your- 
selves, What have I done for my education ? then, as you 
advance in life, What have I done for my country ? so that 
some day that supreme happiness may come to you, the con- 
sciousness of having contributed in some manner to the prog- 
ress and welfare of humanity. 



PREFACE 

The main object of this handbook is to give a concise 
account of the chief chemical antiseptics which have been 
found useful for surgical purposes during the present war. 
Some of the publications on this subject are not readily 
accessible to many who wish to inform themselves as to cur- 
rent European practice, and the requisite information has 
not yet, so far as we know, been collected into a form handy 
for reference. It appeared, therefore, that the present 
work might prove of use to surgeons and others in this 
country who are now taking up military duties connected 
with the care of the wounded. 

The unparalleled severity and frequency of wound in- 
fections in the present war has led to considerable advances 
in our knowledge of antiseptics and of methods for their 
successful employment. These advances have already 
proved to be of great value in the treatment of septic condi- 
tions in civil and industrial practice. 

No endeavor has been made to make a complete com- 
pendium of the innumerable antiseptics and disinfectants 
that have been proposed from time to time, for text-books 
already exist in which most of these substances are ade- 
quately described, and no good purpose would be served by 
duplication. Our principal aim has been to collect in con- 
venient form the methods of preparation and use of various 
new antiseptics and modifications of old ones which have 
received some measure of endorsement by military surgeons 
during the past three years. 



viii PREFACE 

The utility of antiseptics, properly used as adjuncts to 
efficient surgery, is becoming more firmly established as the 
war proceeds. Surgeon General Sir George Makins in the 
Hunterian Oration for 191 7 says: "The most useful practi- 
cal test of the efficiency of any method of wound treatment 
is furnished by the observation of the dates at which micro- 
organisms disappear from the surface of the exposed tissues 
and at which the wound may be safely and permanently 
closed by suture or other means. When subjected to this 
test the antiseptic method has proved itself more rapid and 
more trustworthy than the phylacagogic or saline method 
of treatment." 

A large part of the investigations on antiseptics carried on 
during the last three years has been done at the instance 
of the British Medical Research Committee. We are in- 
debted to Dr. W. Morley Fletcher of that Committee and 
to the editors of the British Medical Journal and the Journal 
of the Royal Army Medical Corps for permission to use parts 
of the reports published in the journals referred to. 

Purely surgical details concerning the use of antiseptics 
do not lie within the scope of the present book although some 
of the principles essential to the successful use of antiseptics 
are briefly referred to. A short statement of the use of 
antiseptics in the treatment of carriers of infectious organ- 
isms is, however, included. Owing to similarity in the 
substances used we have deemed it advisable to include 
a brief statement of the use of certain disinfectants of the 
chlorine group for the sterilization of drinking water and 
the disinfection of hospital ships. 

September 1, 191 7. 

The Herter Laboratory 

New York. 



CONTENTS 



PAGE 

Preface . . . . ' vii 

Chapter I. General Introduction : Classification — laws of 
disinfection — influence of media — choice of 
antiseptics — modes of application . . i 

Chapter II. Antiseptics of the Chlorine Group : Hypochlorous 
acid and hypochlorites, eupad, eusol, chlora- 
mine-T, dichloramine-T, chemical determina- 
tion of the strength of chlorine antiseptics . 17 

Chapter III. The Phenolic Group of Antiseptics : Phenol, cre- 
sols, ]ysol, thymol, salicylic acid, /3-naphthol, 
picric acid, etc 43 

Chapter IV. Salts of the Heavy Metals as Antiseptics : Mer- 
cury, silver, bismuth and zinc salts . . 50 

Chapter V. Dyes as Antiseptics : [Malachite green, brilliant 

green, acriflavine, proflavine . . . .61 

Chapter VI. Miscellaneous Antiseptics : Peroxides, ozone, 
iodine, boric acid and salts, persulphates, acids, 
alcohol, ether, formaldehyde, hexamethylene- 
tetramine, iodoform, permanganates, quinine, 
chinosol, acetanilide 68 

Chapter VII. Methods of Testing Antiseptics : Lethal concen- 
trations — influence of media — time relations 

— velocity — results 77 

Chapter VIII. Certain Special Applications of Antiseptics : Dis- 
infection of carriers — disinfection of water 

— disinfection of hospital ships, etc., with 
electrolytic hypochlorite .... 99 



A HANDBOOK OF ANTISEPTICS 

CHAPTER I 
GENERAL INTRODUCTION 

The terms " antiseptic," " disinfectant/' and " germicide" 
are frequently used irrespective of their precise significance. 
Strictly speaking, an antiseptic is a substance which inhibits 
the reproduction of microorganisms, but it need not of neces- 
sity manifest great killing or "germicidal" action on such 
organisms. Substances such as boric acid or sodium benzoate 
are examples of compounds which are fairly effective in re- 
straining the multiplication of bacteria though possessing 
feeble germicidal properties. Most of the substances used 
in the prevention of wound sepsis possess both antiseptic 
and germicidal properties though perhaps they are most 
commonly termed antiseptics. 

The term " disinfectant " should clearly connote a substance 
which destroys infecting agents and hence is identical with 
a germicide, but the word has come to be used in a popular 
sense, irrespective of complete sterilization, to indicate 
some of the phenomena commonly associated with efficient 
disinfection, such as a deodorant effect. 1 The use of the 
words " disinfectant " and " disinfection " in any other sense 
than that first indicated is to be deprecated. 

1 An account of many important hygienic applications of disinfectants which 
are beyond the scope of the present work will be found in Colonel Lelean's book 
on "Sanitation in War." Churchill. London. 



2 A HANDBOOK OF ANTISEPTICS 

Classification of Antiseptics. — For various reasons it 
is quite impossible to formulate a perfectly logical classifica- 
tion of antiseptics. In the first place, almost every soluble 
substance, provided it can be obtained in sufficient concen- 
tration, is capable of exerting some antiseptic action, so 
that theoretically it would be necessary to classify most 
known organic and inorganic substances, and such an un- 
dertaking is obviously useless and unnecessary. For prac- 
tical considerations it is sufficient to limit ourselves to 
those substances which have found fairly extended use as 
antiseptics. In classifying these we may endeavor to di- 
vide them according to the nature of the substances them- 
selves or according to their mode of action. The latter 
method has much to commend it, but unfortunately we 
know but little of the intimate details of the act of disinfec- 
tion as carried out with even the simplest substances, such 
as phenol or hydrogen peroxide. 

Chemical antiseptics naturally fall into two main classes 
of compounds — inorganic substances as exemplified by 
bodies such as hydrogen peroxide, chlorine, and metallic 
salts, such as those of mercury and silver, on the one hand, 
and on the other organic, or carbon, compounds such as 
phenols, aromatic acids, dye-stuffs, such as malachite green, 
etc., etc. But in these two main groups of inorganic and 
organic antiseptics are found substances of widely different 
properties and above all showing enormous variations in sta- 
bility. This instability of some of the most widely used anti- 
septics is a point of fundamental importance, for upon a clear 
recognition of this fact will depend, to a considerable extent, 
success or failure in their practical use. This chemical insta- 
bility is of various kinds, as a few examples will serve to show. 

Hydrogen peroxide is a good example of an unstable in- 
organic disinfectant. The exact way in which it destroys 
microorganisms is not clear but is undoubtedly connected 
with its oxidizing action. Now when hydrogen peroxide is 



GENERAL INTRODUCTION 3 

placed on the surface of an infected wound, it is rapidly de- 
composed, part of the oxygen of the peroxide being used up 
in oxidizing organic compounds and much of it being liber- 
ated as oxygen gas owing to the decomposition of the peroxide 
by a widely distributed enzyme, "catalase." The net 
result is that after a period of time which may be almost 
incredibly short no undecomposed hydrogen peroxide re- 
mains and disinfection proceeds no further towards comple- 
tion. For, as will be shown later, the rate at which disinfec- 
tion proceeds is directly related to the concentration of the 
antiseptic. Obviously, this instability indicates that hydrogen 
peroxide should be either frequently renewed or restricted in 
its use to conditions where temporary action only is required. 

The chlorine group of antiseptics includes a number of 
important substances such as chlorine itself, hypochlorous 
acid and its sodium and calcium salts, and organic "chlor- 
amines" — i.e. substances containing chlorine attached to 
nitrogen in the form of XC1 groups. They are all char- 
acterized by marked instability, since in disinfection they 
react not only with the cell constituents of microorganisms 
but also with most other substances which are apt to ac- 
company bacteria. In so reacting, the active chlorine of 
the antiseptic is eventually converted either into inert chlo- 
rides or into inert organic substances in which the chlorine has 
become united to carbon. Thus in using the chlorine anti- 
septics, as with hydrogen peroxide, the process of disinfec- 
tion will only go on so long as some of the active substance 
remains undecomposed, and in practical use means must be 
taken for the renewal of these antiseptics at suitable inter- 
vals, in cases where sterilization is not promptly completed. 

In the examples of unstable antiseptics just cited, the 
active substances are decomposed during disinfection and 
cannot be regenerated. In another large class of antiseptics 
the active agent is not totally destroyed during disinfection 
but is rendered relatively inactive. Good examples of this 



4 A HANDBOOK OF ANTISEPTICS 

are found among many metallic salts; mercuric chloride 
and silver nitrate, for example, are much less effective when 
acting on bacteria suspended in hard water than when acting 
on washed bacteria suspended in distilled water. Similarly, 
phosphates and other salts frequently found in wound exu- 
dates inhibit the action of metallic antiseptics. In these 
cases disinfection by the metallic salts is impeded not by 
the complete destruction of the compound, as with hydrogen 
peroxide or chlorine antiseptics, but through the conversion 
of the metallic salt into an inactive form. This conversion 
may be effected either by precipitation of the metallic com- 
ponent in an insoluble form, or by changing its state — e.g. 
changing its condition of ionization in aqueous solution. In 
such cases the antiseptic substance is rendered inert but not 
destroyed and, theoretically at least, could be regenerated 
by suitable chemical means. 

Lastly, there are other substances used as antiseptics which 
apparently are stable during disinfection or at least com- 
paratively so. Familiar examples of compounds of this 
type are found in the phenols and in aromatic substances of 
the dye class. These compounds apparently are not readily 
decomposed by the cell constituents of bacteria nor by 
most substances which are apt to accompany the latter. 
It has been suggested that some of these antiseptics act by 
virtue of changes they produce in the surface layer of the 
bacterium so that the latter is rendered permeable by the 
antiseptic. But even with these relatively stable substances 
there is a definite limit to the amount of disinfection which 
any given quantity of the compound can accomplish and 
this amount will vary according to the conditions under 
which the antiseptic acts. This fact will be noted again 
in the following section in which some reference is made to 
the laws governing disinfection. 

The following table shows the relative stability or insta- 
bility of some of the common groups of antiseptics : 



GENERAL INTRODUCTION 



Inorganic Antiseptics 

Hydrogen Peroxide and some of its 

derivatives 
Chlorine 



Hypochlorous Acid and its Salts 

Bromine and Iodine 
Boric Acid and its Salts 
Mercury Salts 

Silver Salts 

Bismuth Salts 

Zinc Salts 



Unstable, easily decomposed during 

disinfection 
Unstable, easily decomposed during 

disinfection 
Unstable, easily decomposed during 

disinfection 
Less unstable than chlorine 
Stable 
Often inactivated by precipitation or 

otherwise 
Often inactivated by precipitation or 

otherwise 
Often inactivated by precipitation or 

otherwise 
Often inactivated by precipitation or 

otherwise 



Alcohol, Ether, etc 
Iodoform 
Formaldehyde 

Hexamethylenetetramine and its de- 
rivatives 
Phenols, Naphthols, and derivatives 
Aromatic Chloramines 

Dyes such as malachite green, acri- 
flavine, etc. 



Organic Antiseptics 
Stable 



Fairly stable 
Unstable 
Mostly stable 

Mostly stable 

Unstable, easily decomposed during 
disinfection 

Mostly stable, though sometimes re- 
duced to leuco-forms, and often 
adsorbed by tissues 

It is a surprising fact that this varying stability of anti- 
septics is so little realized by many workers. Current litera- 
ture is full of reports of experiments in which bacteriologists 
have added reactive unstable substances, such as hypo- 
chlorous acid, to media which cause their prompt disap- 
pearance in dilute solution ; the mixture containing little or 
no remaining antiseptic is subsequently sown with organ- 
isms and astonishment is expressed at the inefficiency of the 
substance employed as a disinfectant. This important point 
will be referred to repeatedly in later sections. 

The Laws Governing Disinfection. — These have been 
chiefly worked at by Miss Chick. 1 Earlier experiments were 

1 Journal of Hygiene, 8, p. 92, 1908, and 10, p. 238, 1910. 



6 A HANDBOOK OF ANTISEPTICS 

carried out by Kronig and Paul 1 and by Madsen and Nyman. 2 
It would be out of place to go into the details of the elaborate 
experiments and calculations necessary to elucidate the 
principles of disinfection but a brief summary of the essential 
results may be given. Miss Chick observed the number of 
bacteria that survived, at varying intervals of time, the 
action of a constant quantity of a stable antiseptic, such as 
phenol, at a constant temperature, on a known number of 
organisms. By utilizing these results mathematically she 
was able to calculate a velocity coefficient for the disinfecting 
action of the substance. The main result of these experi- 
ments was to show that in all essential particulars the act 
of disinfection could be regarded as obeying the laws govern- 
ing a simple chemical reaction, the disinfectant representing 
one reagent and the bacteria the other. This conception is 
of the greatest importance since the cardinal points of 
efficient disinfection, namely adequate active mass or con- 
centration of antiseptic, time of action, and perfect contact, 
are thereby experimentally established. 

The influence of temperature upon the rate of disinfection 
is interesting since here again a close analogy exists be- 
tween ordinary chemical reactions and disinfection. The 
velocity of disinfection increases with rise in temperature in 
a manner similar to that of an ordinary chemical reaction. 
Some idea of the magnitude of the effect of temperature 
may be gathered from the fact that with metallic salts the 
mean velocity of disinfection increased 2-4 fold for a rise 
in temperature of io° C, while with phenol it was as high as 
eightfold, using B. paratyphosus as test organism in each case. 

It must be remembered, however, that the similarity in 
the mathematical relations governing the velocity of simple 
chemical reactions and disinfection does not of necessity prove 
that the act of disinfection is a chemical one, although with 
some antiseptics this is almost certainly the case. 

1 Zeitschr. f. Hygiene, 25, p. 1, 1897. 2 Ibid., 57, P- 388, 1907 






GENERAL INTRODUCTION 7 

Influence of Media. — The capacity of a disinfectant 
to kill microorganisms is dependent to an extraordinary de- 
gree upon the conditions under which it acts. Almost in- 
variably the greatest germicidal activity is shown when the 
substance acts upon bacteria freed from contaminating cul- 
ture media and suspended in distilled water or salt solution. 
The presence of proteins and similar substances, e.g. peptones, 
usually causes a huge reduction in the germicidal activity 
of most substances. In the presence of pus, in which many 
of the organisms are partly ingested in the bodies of the 
dead leucocytes and hence difficult to attack, the results are 
still less favorable. The following table gives some idea 
of the reduction in germicidal activity of some of the common 
antiseptics acting for two hours at room temperature on 
staphylococcus aureus (i drop of 24 hours broth culture) in 
a total volume of 5 cc. of either water or 50 per cent horse 
serum. The antiseptics were in all cases added last of all 
to the mixtures : 



Antiseptic 



Phenol .... 
Salicylic Acid . . 
Hydrogen Peroxide 
Iodine .... 

Mercuric Chloride 1 
Silver Nitrate x . . 
Sodium Hypochlorite 
Chloramine-T . . 



Staphylococci 
in Water 

' 1 : 250 - 
, 1 : 500 + 

1 : 2500 — 
k 1 : 5000 + 
' 1 : 3500 - 
k 1 : 8000 + 

1 : 100,000 — 
L 1 : 1,000,000 + 

1 : 5,000,000 — 
> 1 : 10,000,000 + 

1 : 1,000,000 — 
k 1 : 10,000,000 + 

1 : 500,000 — 
. 1 : 1,000,000 + 
' 1 : 500,000 — 

1 : 1,000,000 + 



Staphylococci in 50% 
Horse Serum 
1:50 - 
1 : 100 + 
1 : 100 — 
1 : 250 + 
1 : 1700 — 
1 : 2000 + 
1 : 1000 — 
1 : 2500 + 
1 : 25,000 — 
1 : 50,000 + 
1 : 10,000 — 
1 : 25,000 + 
1 : 1500 - 
1 : 2000 + 
1 : 2000 — 
1 : 3000 + 

negative subcultures. 



The — sign indicates sterilization as evidenced by 
while the + sign indicates incomplete disinfection. 

1 No antidote was used in these experiments to prevent the action of traces of 
the salt in the subcultures. (Cp. p. 52.) 



8 A HANDBOOK OF ANTISEPTICS 

In order to illustrate the difficulty of sterilizing pus, some 
selected experiments made by W. Parry Morgan x may be 
cited. In these experiments one part of pus was mixed with 
nine parts of antiseptic and after ten minutes 10 cmm. were 
planted in liquid agar which was then shaken and incubated. 
It was found that when the antiseptic was strong the num- 
ber of colonies could be counted readily, but when it was 
weak the agar became opaque with innumerable colonies. 
The results are, of course, comparative and do not lend 
themselves to strict quantitative expression, for the specimens 
of pus vary much among themselves. 

Table showing the Effects of Antiseptics upon the Growth 
of Bacteria in Pus 

Iodine i : 400 1 : 800 1 : 1600 1 : 3200 

16 hours -f ++ + + ++ + + 

Mercury Biniodide 1 : 400 1 : 800 1 : 1600 1 : 3200 

16 hours + ++ ++ + + + 

Phenol 1 : 40 1 : 80 1 : 160 

16 hours . ; + ++ + + + 

Sodium Hypochlorite 1 : 200 1 : 400 1 : 800 1 : 1600 

(Dakin's Solution) 

18 hours — — — + 

3 days — - - + 

Eusol 

(Bleaching Powder and Boric Acid) 

available chlorine 1 : 200 1 : 400 1 : 800 1 : 1600 

18 hours — — — + 

3 days ........ — - - + + 

When the proportion of pus to antiseptic was higher than in 
the above experiments, the disinfecting action was still fur- 
ther diminished. 

A few substances, such as some derivatives of hexameth- 
ylenetetramine, are known whose germicidal action is said 
not to be materially affected by the presence of serum, and 
one striking example exists of a substance which is apparently 
actually more active in serum than in water. This compound 
prepared by Benda and termed trypaflavine but now known 

1 Brit. Med. Journ., May 13, 1916. 



GENERAL INTRODUCTION 9 

as acriflavine (sometimes simply flavine) has been shown by 
Browning l and his colleagues to kill staphylococci in serum 
at one tenth the concentration necessary when the same or- 
ganisms are suspended in peptone solution. Experiments 
with pus, however, showed much higher concentrations were 
necessary to achieve sterilization than with peptone or serum 
media. 

The causes of these huge variations in the germicidal ac- 
tion of antiseptics under various conditions are but slightly 
understood. The reduction in the case of members of the 
chlorine group can be accounted for to a considerable extent 
by the rapidity of the chemical interaction of the disinfectant 
with the protein medium, with the production of inert 
substances if the proteins be in sufficient excess. But other 
causes must be at work where the more stable antiseptics 
such as phenol and the dye-stuffs are concerned. It is 
frequently said that the antiseptic is " quenched " or " fixed " 
by the protein medium, but these terms do not carry much 
enlightenment, nor does the suggestion that the molecular 
condition of the antiseptic is influenced by the presence of 
colloids in the medium. The low germicidal action shown by 
most antiseptics against pus is due in part no doubt to the 
mechanical difficulties of penetrating the mucoid particles 
in the pus. Parry Morgan has shown, moreover, that when 
the organisms in pus, or added to it, have undergone phago- 
cytosis they are less readily destroyed by antiseptics. The 
reduction in antiseptic activity of a substance in the presence 
of any particulate matter has been often observed 2 and is 
probably connected with the surface adsorption of the anti- 
septic by the particles with consequent reduction in its ef- 
fective concentration. 

The Choice oe Antiseptics. — The selection of anti- 
septics for various purposes requires the consideration of a 

1 Brit. Med. Journ., Jan. 20, 1917. 

2 Chick and Martin, Journ. of Hygiene, 8, p. 654, 1908, 



10 A HANDBOOK OF ANTISEPTICS 

number of factors. The possession of high germicidal ac- 
tivity, as ordinarily tested in the presence of media comparable 
with those in which the disinfectant is to act, is of course 
desirable. But it is equally important to bear in mind the 
concentration at which the substance may be used, for a 
relatively innocuous substance used in fairly high concentra- 
tion will often give much more satisfactory results than lower 
concentrations of more active substances. The speed of 
disinfection is also an important question for it varies enor- 
mously with different types of antiseptics. Antiseptics of 
the chlorine group and iodine are among the most rapid, 
while the dyes and some metallic salts are relatively slow. 
An inspection of the tables in Chapter VII, in which a heavily 
infected mixture of blood serum and muscle extract was 
treated with various antiseptics at about the concentration 
recommended for wound treatment will give a good idea of 
their relative speed of action and potency under the conditions 
selected. 

The ideal surgical antiseptic should effect complete steri- 
lization within its sphere of action without causing any 
damage to animal cells. At the moment such a substance 
does not appear likely to be found, but on the other hand 
it is surprising to see how little damage may be done to 
animal tissues by some active antiseptics. An important 
method of judging of the injurious action of antiseptics is to 
investigate the condition of the leucocytes in wounds re- 
cently treated with the substance under consideration. In 
general it appears from experiments in vitro that, with the 
strength of antiseptics commonly used in surgery, mercury 
salts and hypochlorites have relatively little effect on phago- 
cytosis as compared with phenol (Parry Morgan). It is a 
regular phenomenon to observe activity of the leucocytes 
obtained from wounds which have been recently treated 
with hypochlorites. 

Ingenious methods for determining the influence in vivo 



GENERAL INTRODUCTION 11 

of antiseptics on the activities of leucocytes have been worked 
out by Col. C. J. Bond. 1 Indigo impregnated threads are 
laid in aseptic or septic wounds with or without antiseptics. 
After varying periods of time the threads will be found to 
have become decolorized owing to the ingestion of pig- 
ment particles by the leucocytes. It was found that the 
application of a strong antiseptic, e.g. i : iooo mercury 
biniodide or i : 20 phenol, to a wound such as that made for 
the radical cure of hernia or for the removal of varicose veins, 
does delay to some slight extent the decolorization of an 
indigo thread placed in it as a drain. But in general it ap- 
peared that antiseptic solutions in moderate concentration 
exercise less influence over emigration and phagocytosis than 
many surgeons had supposed. There are, however, rea- 
sons for concluding that many antiseptics do exert a con- 
siderable inhibitive effect on the return immigration of liv- 
ing phagocytes. But if the use of antiseptics does bring 
about a reduction of the numbers, or inhibit the activities, 
of pathogenic organisms, not merely in vitro but in the 
actual wound, then although these reagents undoubtedly do 
cause the death of a certain number of body cells and prevent 
others from again reaching the tissues, this is relatively 
of slight importance if the invading organisms are at the 
same time materially diminished in numbers or offensive 
capacity. 

Other criteria which need consideration in judging of the 
suitability of any particular substance as an antiseptic con- 
cern the absence of marked irritation of the skin or other tis- 
sues to which the substance may be applied, and also its 
effect on the rate of dissolution of necrotic tissue and the 
rate of formation of healthy granulations. In general, too, 
it will be found that antiseptics which coagulate the proteins 
in wound exudates, e.g. salicylic acid, or strong phenol, are 
less desirable than those which do not act in this way, 

1 Brit. Med. Journ., June 3, 1916, Feb. 3, 191 7. 



12 A HANDBOOK OF ANTISEPTICS 

since anaerobes are apt to multiply in necrotic tissue more or 
less surrounded with impermeable coagulum. In contrast 
with this effect, the solvent action of hypochlorites and the 
related dichloramine-T on dead tissue is marked and con- 
stitutes a valuable property. 

But the problem of selecting the most desirable antiseptic 
is by no means limited to the preceding considerations. 
It is of paramount importance that judgment should be 
exercised in choosing a substance which is likely to be effec- 
tive under the conditions of its employment. Thus any of 
the hypochlorite solutions which give excellent results when 
used for the intermittent flushing of infected wounds, partly 
on account of the rapidity of their action and of their gen- 
erally inoffensive character, are almost useless when ap- 
plied in small amounts as a simple wet dressing which is 
infrequently renewed. In the latter case no active anti- 
septic persists for any significant length of time and not much 
more is accomplished than the prevention of secondary rein- 
fection. When prolonged action is required and frequent 
application is impossible, recourse must be had to more 
stable mixtures which yield up their store of antiseptic slowly. 
Examples of such substances are the bismuth iodoform par- 
affin mixture (B.I. P.) and the oi]y solution of dichloramine- 
T. These substances will be referred to later. 

Finally, it must be once more emphasized that antiseptics 
at best are only accessories to, and not substitutes for, efficient 
surgical treatment of infected wounds. The disinfecting 
action of antiseptics is practically restricted to the wound 
surfaces and the cavity inclosed by them and none of them 
appears to have any considerable power of penetration with- 
out simultaneous loss of activity. The prompt excision of 
damaged tissue has now become a routine method in war 
surgery on the Western Front, and this procedure, renders 
the effective employment of antiseptics much more certain 
than it was earlier in the war. 



GENERAL INTRODUCTION 13 

Modes of Application. — Antiseptics are applied in 
various ways, according to the character of the wound, the 
nature of the substance, and the kind of action desired. 
Aqueous solutions are used most extensively. When un- 
stable antiseptics, such as hypochlorites and chloramine-T, 
are used and prolonged antiseptic action is required — as 
in the treatment of freshly infected or septic wounds — it is 
necessary to employ some means of frequently renewing the 
antiseptic. This end is- commonly accomplished either by 
irrigation or by intermittent instillation of fresh antiseptic 
solution into the wound by means of short rubber tubes so 
arranged that the solution may reach every recess of the 
wound. This method has been used extensively in the pres- 
ent war and has given excellent results. The technique of 
the method will be found fully described in a book by Carrel 
and Dehelly. 1 To obtain the best results with this mode of 
treatment, a careful observation of details is essential, and 
as only an outline can be given here reference should be 
made to the book just mentioned or to numerous original 
papers in recent publications. 

The first step is the mechanical cleansing of the wound 
by excision, this being omitted as dangerous if the phenomena 
of inflammation have already set in. The wound is then 
subjected to active antiseptic treatment. For this purpose 
sodium hypochlorite — so-called Dakin's solution — is em- 
ployed at a concentration of 0.5 per cent (see page 23). 
Great care is taken to insure uniform distribution of 
the antiseptic by means of the rubber tubes w T hich are 
usually of fine bore, closed at one end and provided 
near the closed end with a number of fine lateral per- 
forations. A number of these tubes, depending on the 
size of the cavity, are disposed within the wound, care being 
taken that no gauze intervenes between the tubes and the 

1 Le traitement des plaies infectes, Collection Horizon, Masson et Cie., 
Paris, 191 7, 



14 A HANDBOOK OF ANTISEPTICS 

tissues and also that the tubes are not so closely applied 
as to block the perforations. The tubes are led through the 
outer dressing and kept in position with safety pins. When, 
as is always the case with large wounds, more antiseptic 
solution is needed than can be conveniently supplied by using 
a glass syringe, the ends of the tubes are connected by T 
pieces to a single tube which in turn connects with a glass 
reservoir holding a liter of solution which is suspended sev- 
eral feet above the level of the bed. The supply of solution 
to the wound is arranged by opening for a few seconds 
every two hours the clip attached to the main tube. An 
important point is the absence of a drainage opening at the 
most dependent part of the wound; in fact, the ideal ar- 
rangement is attained when the wound is cup-shaped; 
when it is on the inferior surface of a limb the aperture is 
partly plugged with gauze to hinder free escape of the fluid. 
It is important to note that vessels in the base of the wound 
should not be ligatured with silk since this material is at- 
tacked by hypochlorites. The routine bacteriological ex- 
amination of the wound is important. It is found that when 
practical sterility has been maintained for two or three 
days it is safe to close the wound. In the case of wounds 
that have suppurated, it is advisable to wait a little longer. 

In place of the hypochlorite, the more stable chloramine- 
T has been used to a considerable extent and with particular 
success in fresh industrial accidents. It is usually used in 
0.5-2 per cent solution and its action is similar to that of 
hypochlorite save that necrotic tissue is less rapidly removed. 

The methods of application of the ordinary solutions of 
less unstable antiseptics such as phenol, iodine, and the 
metallic salts, call for no special comment. 

When the frequent renewal of the antiseptic is impracti- 
cable or not desired, recourse may be had to pastes or 
oils of various kinds, which embody a store of antiseptic 
that may be gradually utilized. The mild antiseptic action 



GENERAL INTRODUCTION 15 

of a paste containing about one per cent of chloramine-T 
and five per cent of sodium stearate in water is often 
useful to maintain sterility and prevent reinfection of 
wounds which have previously received more vigorous 
treatment. A much more intensive action can be obtained 
from the use of dichloramine-T dissolved in eucalyptol 
and paraffin as described on page 39. The amount of 
active antiseptic which may thus be applied to the wound 
surface is extremely large and its action is correspondingly 
prolonged. The method of using this oil is particularly 
simple and the results appear to be exceptionally good, 
both as regards the prevention and treatment of sepsis. 

A paste possessing moderate potency, introduced by 
Rutherford Morison, composed of bismuth subnitrate, 
iodoform, and paraffin, is being widely used with good results 
both for wounds of the soft tissues and for fractures. The 
fact that the wounds do not need dressing for several days 
gives it a great advantage when the number of cases re- 
quiring treatment is very large, and because of this it also 
secures a maximum of rest for the wound. 

It must not be forgotten that the employment of pastes 
as a primary dressing is not free from serious risks and 
should only be practiced with caution. The use of salicylic 
acid pastes for the immediate treatment of wounds other 
than superficial ones is said to have given particularly poor 
results as anaerobes are apt to flourish in walled off recesses 
of the wound. The coagulating properties of salicylic acid 
would especially facilitate such a result. 

The administration of antiseptics by intravenous injection 
is the only method other than that of local application that 
is at all practiced for military purposes. Eusol has been 
used to some extent for this purpose l though it is abundantly 
clear that any action it may have is unconnected with any 

1 Lorrain Smith, Ritchie, and Rettie, Brit. Med. Journ., Nov. 13, 191 5. Frazer 
and Bates, Brit. Med. Journ., Aug. 5, 1916. 



16 A HANDBOOK OF ANTISEPTICS 

germicidal effect since the actively haemolytic hypochlorite, 
in therapeutic doses, is immediately decomposed by the 
blood. 1 A few striking cases of favorable results of its use 
have been recorded but as the procedure has not found wide 
acceptance and is not free from danger, it is well to suspend 
judgment as to its value. A number of years ago 2 silver 
nitrate was used intravenously in the same way and while it 
was clear that no direct germicidal effect was possible, some 
kind of beneficial effect was believed to follow the haemolysis 
caused by the silver salt. 

1 Dakin, Brit. Med. Journ., June 17, 1916. 

2 Klinisch-Therapeutische Wochenschrift, No. 33, p. 881, 1908, 



CHAPTER II 
ANTISEPTICS OF THE CHLORINE GROUP 

In the present war, which is distinguished by the frequency 
and intensity of virulent wound infections, no class of anti- 
septics has received such extensive employment as those of 
the chlorine group. When properly applied — and this is 
an important reservation — it is generally conceded that 
they have proved of genuine value to the surgeon. All of 
the antiseptics of this group are characterized by chemical 
instability in the presence of organic matter, and therefore 
conditions favorable for their use must include either pro- 
vision for their frequent renewal, or the use of some im- 
miscible solvent for the antiseptic so that the active compound 
may be gradually liberated. 

The members of the group under consideration are often 
spoken of as containing " active" chlorine as distinct from 
inert chlorine such as that in common salt. It must not be 
inferred, however, that the phrase " active chlorine" neces- 
sarily implies either that free chlorine is contained in the 
substance or is liberated from it, as more often it will be 
found that hypochlorous acid or some such compound is 
the active agent. From the standpoint of disinfection, it 
is probably correct to connote with the term " active chlorine " 
in a compound, the ability of any particular substance to 
part with chlorine, free or combined, in such a way that it 
can effect the chlorination of bacterial and other proteins. 1 

1 All compounds containing active chlorine possess the property of liberating 
iodine from an acidified solution of potassium iodide. The iodine may be de- 
tected by the development of a blue color on adding starch paste. This reaction 
will often be found useful in testing for the presence of unchanged antiseptic. 
C 17 



18 A HANDBOOK OF ANTISEPTICS 

This chlorination of bacterial protein seems to be incompatible 
with the life of the microorganism. It is necessary to refer 
in outline to the nature of this reaction. All proteins, irre- 
spective of their origin, contain large numbers of amino-acid 
groups, which may be represented as shown below : (I) 

H H 

I I 

R— C— CO R— C— CO— 

I I 

NH NCI 



I II 

These groups are capable of attack by substances containing 
active chlorine in such a way that the hydrogen attached to 
the nitrogen atom is replaced by chlorine (II). The com- 
pounds thus formed contain the (NCI) group and hence 
belong to the class of chloramines. Their chlorine is still 
active and they are themselves active germicides. Other 
concomitant reactions also occur which use up part of the 
chlorine, converting it into an inert form, e.g. the chlorine 
becomes united to carbon or forms chlorides. 

This formation of germicidally active chloramines is of 
importance in several respects. For example, the proteins 
and other nitrogenous compounds present in wound secre- 
tions may be converted into chlorine derivatives of antiseptic 
value by the action of a sufficient quantity of hypochlorite 
or similar substance. 1 While preformed chlorine derivatives 

1 It is of interest to note that while chlorine, bromine, and iodine have not 
widely differing germicidal properties, hypobromites and hypoiodites in contrast 
to hypochlorites have but trivial disinfecting action. Correlated with this fact 
is the observation that hypochlorites react readily with proteins while hypo- 
bromites and hypoiodites do not. On the other hand, certain synthetic broma- 
mines which react readily with amino-acids and proteins have a high germicidal 
potency. The ability to react with proteins and allied bodies is clearly associated 
with germicidal activity in members of the halogen group of antiseptics. 



ANTISEPTICS OF THE CHLORINE GROUP 19 

prepared from proteins are powerful germicides, they are 
not convenient substances for general use, but by using 
other types of nitrogen compounds for chlorinating, synthetic 
chloramines, with valuable antiseptic properties, are readily 
obtainable. 

For detailed information concerning the action of chlorine 
antiseptics upon amino-acids, proteins, etc., as well as the 
preparation of many synthetic chloramines, reference must 
be made to the original papers. 1 

In addition to their disinfecting action, the chlorine 
antiseptics are strong oxidizing agents and deodorants and 
moreover possess in high degree the property of decompos- 
ing toxins. By the regulated action of hypochlorous acid, 
Dean 2 has prepared a non-toxic dysentery vaccine and it 
is a common observation that the free use of hypochlorites 
may reduce the constitutional symptoms arising from septic 
processes and that they reappear on discontinuing the anti- 
septic treatment. 

The antiseptics of the chlorine group which are most com- 
monly employed in the treatment of infected wounds, are the 
following : 3 

(a) Hypochlorous acid and its sodium and other salts 
(including "eupad," "eusol" and so-called Dakin's solution). 

(b) Chloramine-T, the abbreviated name for sodium 
toluene sulphonchloramide. 

(c) Dichloramine-T, the abbreviated name for toluene 
sulphondichloramine. 

In most respects, the action of these various chlorine 
compounds is essentially similar, though each possesses 
certain properties which render it more or less suitable for 

1 Brit. Med. Journ., Jan. 29, 1916, June 17, 1916; Proc. Roy. Soc. B, 8q, 
p. 232, 1916. Biochem. Journ., June, 1917. 

2 Brit. Med. Journ., April 29, 1916. 

3 Chlorine water has been used to some extent, but it is doubtful whether its 
use is preferable to that of the more convenient iodine solution, which in most 
respects it resembles. 



20 A HANDBOOK OF ANTISEPTICS 

particular purposes. As a matter of convenience, it may 
be desirable to give a short resume of these considerations. 

I. Hypochlorous acid and hypochlorites are best suited to 
cleansing septic wounds by irrigation. They markedly 
assist in the dissolution of necrosed tissue. They are unstable 
and very reactive, and must be frequently renewed to all 
parts of a wound, this being best achieved by the method of 
intermittent instillation (p. 13). They are the cheapest 
antiseptics of the chlorine group and are much cheaper than 
other effective germicidal substances. Free hypochlorous 
acid is more irritating than the sodium salt. The latter in 
0.5 per cent neutral or feebly acid (boric acid) solution may 
be used in large quantities under appropriate precautions 
for a considerable time without causing irritation. The 
skin is more susceptible than the deeper tissues and should 
be protected with vaseline or some similar substance. 

II. Chloramine-T can be used in stronger solution (up to 
two per cent) than the hypochlorites. It is more stable 
and exerts more prolonged antiseptic action and is consider- 
ably more effective than hypochlorite when acting in the 
presence of much blood. It is not toxic and is less irritating 
than the hypochlorites and has but little solvent action on 
necrosed tissue. It is well suited for use on wounds previously 
cleansed with hypochlorites or dichloramine-T, and in suitably 
dilute solutions may be used in the eye and on other sensitive 
parts. It may be applied in solution, as an impregnation 
of gauze, or in a sodium stearate cream. 

III. Dichloramine-T dissolved in oily media may be 
sprayed upon wound-surfaces or poured into accessible 
parts of deep wounds. It yields moderate amounts of anti- 
septic to watery media such as secretions from wounds or 
mucous membranes. It is suitable for cases requiring pro- 
longed antiseptic treatment, and for first dressings of recent 
wounds which do not require irrigation. It is also used for 
nasal antisepsis (p. 102). Dichloramine-T in oil solution has a 



ANTISEPTICS OF THE CHLORINE GROUP 21 

great advantage over the other chlorine antiseptics in that 
it may be used in high concentration, and its action is of much 
longer duration. The application of the oil is extremely 
simple and it ordinarily need not be renewed more than once 
in 24 hours. 

Hypochlorous Acid and its Salts 

The disinfecting action of these substances has been known 
for over a hundred years and they have received numerous 
hygienic applications. An account of their early history 
and uses will be found in the British Medical Journal, Dec. 
4, 1915. In the early part of the present war, several 
surgeons made use of commercial sodium hypochlorite, "eau 
de Javel," for the treatment of infected wounds, but there 
were many accidents owing to its caustic action, which was 
largely due to the presence of excessive amounts of alkali 
in the solution. Apart from this objectionable property, 
the substance appeared to possess desirable qualities and it 
was for this reason that several workers sought to obtain 
hypochlorite solutions which were less irritating but w T hich 
retained their germicidal properties unchanged. 

These various solutions are all prepared from bleaching 
powder (chloride of lime) which is the most readily accessible 
solid source of hypochlorous acid and its salts. Bleaching 
powder is prepared by the action of chlorine upon slaked 
lime and in most respects behaves like a mixture of calcium 
hypochlorite and calcium chloride. It is of variable com- 
position and slowly decomposes on keeping, especially when 
exposed to air or light (p. 41). In making some of the anti- 
septic solutions, the calcium hypochlorite of the bleaching 
powder is converted into sodium hypochlorite by the action 
of sodium carbonate. In most of them boric acid is added 
to counteract the objectionable alkalinity of ordinary hypo- 
chlorites, and this has also the effect of liberating a certain 



22 A HANDBOOK OF ANTISEPTICS 

amount of free hypochlorous acid, leaving the solution 
neutral or faintly acid. Sodium hypochlorite may also be 
prepared by the electrolysis of sodium chloride solution 
(p. 1 1 6) and by the action of chlorine on sodium carbonate 
or sodium bicarbonate. It will be impossible to describe 
the preparation and uses of all the various hypochlorite 
solutions that have been employed for surgical use and it 
would appear sufficient to give an account of the two that 
seem to be most widely used, namely " eusol " and the so-called 
Dakin solution. All hypochlorite solutions attack metals 
and hence they should not be used for the sterilization of 
instruments. 

Eupad and Eusol. 1 — These are preparations of bleaching 
powder and boric acid, either dry (eupad) or in solution 
(eusol). The former is prepared by intimately mixing equal 
weights of bleaching powder and boric acid, both in fine 
powder. It has been employed as a dusting powder, and in 
strands of gauze for drainage or between layers of moistened 
gauze or lint as a dressing. When moistened, eupad liberates 
hypochlorous acid partly in gaseous form and in variable 
amount and this, if excessive, is liable to prove highly irri- 
tating, so that the quantity of eupad used must be carefully 
controlled. Eupad forms a rather thick, white coagulum 
with wound exudates and this may occasion inconvenience. 
It is used much less extensively than is the aqueous eusol. 
An almost identical mixture was recommended by Vincent 
some years ago while Lumiere advocates a mixture con- 
taining bleaching powder one part with boric acid three 
parts. 

Eusol is prepared in either of two ways : 

(i) Twenty-five grams of eupad are shaken up with one 
liter of water, allowed to stand for a few hours, then filtered 
through cloth or filter paper. 

1 Lorrain Smith, Drennan, Rettie, and Campbell, Brit. Med. Journ,, July 24, 
I9I5- 



ANTISEPTICS OF THE CHLORINE GROUP 23 

(2) To i liter of water add 12.5 grams bleaching powder, 
shake vigorously. Then add 12.5 grams boric acid powder 
and shake again. Allow to stand for some hours, preferably 
overnight, then filter off, and the clear solution is ready for 
use. Eusol prepared in this way from good quality bleaching 
powder contains the equivalent of about 0.27 per cent hypo- 
chlorous acid. 1 The solution gives thick precipitates contain- 
ing calcium with blood or wound exudates and is strongly 
hemolytic. It is frequently spoken of as a solution of 
hypochlorous acid but actually the mixture is alkaline to 
litmus and contains a balanced mixture of calcium hypo- 
chlorite and borate with an undetermined amount of free 
hypochlorous acid. The separate estimation of the latter 
is a difficult problem. A large number of experiments on 
the germicidal action of eusol will be found in Lorrain Smith's 
paper, together with suggestions as to methods of use in the 
treatment of infected wounds. It is recommended by its 
authors for use (a) as a lotion, diluted if necessary, (b) as a 
fomentation, (c) as a wet dressing with gauze without a 
waterproof covering, and (d) as a bath, diluted if necessary. 

The general principles concerning the use of chlorine anti- 
septics as germicides apply equally to eusol and the other 
hypochlorites (cp. pp. 12, 20). 

Neutral Sodium Hypochlorite Solution ("Dakin's 
Solution"). — This preparation is essentially a solution of 
sodium hypochlorite, containing 0.45 to 0.5 per cent NaCIO 
made in such a way that it is, and remains, substantially 
neutral. Ordinarily commercial hypochlorite is very variable 
in composition and commonly contains much free alkali 
and occasionally free chlorine. Such solutions are very 
irritating and should not be used for surgical purposes. The 
original formula 2 for making the neutral solution requires 

1 Through an error in calculation, the composition was first given as 0.54 
per cent. The other analytical figures given in the original paper also contain 
errors. 

2 Comptes rendus, 161, p. 150, 1915, Brit. Med. Journ., Aug. 28, 1915. 



24 A HANDBOOK OF ANTISEPTICS 

the use of boric acid for neutralization. The reason for this 
may perhaps be briefly referred to. It is well known that 
blood and some other body fluids and also certain artificial 
salt solutions containing mixtures of the salts of polybasic 
acids — e.g. phosphoric or carbonic acid — ■ are able to retain 
their essential neutrality even after the addition of limited 
quantities of acid or alkali. This is due to the fact that the 
addition of acid or alkali simply changes the relative pro- 
portion of two or more salts of the polybasic acid present in 
the solution. Such solutions are often referred to as " bal- 
anced " and the salts in them are called " buffer salts/' Uti- 
lizing the same principle and employing the feeble polybasic 
boric acid, a simple balanced hypochlorite mixture was 
prepared which maintains essential neutrality under all 
conditions. It should be understood that the insignificant 
antiseptic action of boric acid has nothing to do with the em- 
ployment of this acid nor is the boric acid employed for the 
purpose of liberating free hypochlorous acid as in Lumiere's 
or Lorrain Smith's preparations. 

Preparation of Neutral Sodium Hypochlorite. — 
One hundred and forty grams of dry sodium carbonate 
(Na 2 C0 3 ) or 400 grams of* the crystallized salt (washing soda) 
are dissolved in 10 liters of tap water, and 200 grams of bleach- 
ing powder containing 24-28 per cent of " available chlorine " 
are added. 1 The mixture is very thoroughly shaken, both to 
make good contact and to render the precipitated calcium 

1 Bleaching powder varies considerably in its available chlorine content, 
though when bought in bulk the fresh product is fairly constant in composition. 
It is advisable to determine the " available chlorine " in each large batch of bleach- 
ing powder purchased, as described on p. 41. Bleaching powder with less than 
23 per cent of available chlorine should be rejected. Exceptional samples may 
contain as high as 35 per cent available chlorine and in such cases it is well to 
reduce correspondingly the ingredients taken in the above formula. For pur- 
poses of rough calculation, one may assume that using 200 grams of bleaching 
powder for 10 liters of solution, the resulting product will contain as much 
sodium hypochlorite as is represented by the available chlorine of the bleaching 
powder divided by 50. Thus 25 per cent " available chlorine " bleaching powder 
will give 0.5 per cent sodium hypochlorite solution. 



ANTISEPTICS OF THE CHLORINE GROUP 25 

carbonate granular and promote its settling. It is then 
allowed to stand quietly and after half an hour the clear liquid 
is siphoned off from the precipitate and filtered through a 
cotton plug or paper. Forty grams of boric acid are added to 
the clear filtrate and the resulting solution is ready for use. 
The boric acid must not be added before filtering but only 
afterwards. The exact strength should be determined from 
time to time, as directed on p. 41. It is important that the 
solution should not be stronger than 0.5 per cent sodium 
hypochlorite or irritation of the skin may be frequent. On 
the other hand, it should not be less than 0.4 per cent or its 
germicidal action is materially diminished. The solution 
should also be tested for neutrality by adding a little of it 
to a trace of solid phenolphthalein suspended in water. No 
red color indicating free alkali should develop or else more 
boric acid must be added ; this is, however, rarely necessary 
with the above proportions. The solution should not be 
kept longer than one week. 1 

Daufresne has shown that it is possible to prepare a satis- 
factory solution of sodium hypochlorite without employing' 
boric acid for neutralization if sodium bicarbonate be used 
in conjunction with sodium carbonate for decomposing the 
bleaching powder. The relative proportions of sodium 
carbonate and bicarbonate required to furnish a neutral 

1 A stronger solution may be prepared by decomposing bleaching powder 
with dry sodium carbonate in the proportion of 150 gms. to 105 gms., dissolved 
in 1 liter of water. The mixture is filtered and a measured portion of it (20 cc.) 
rapidly titrated with a boric acid solution of known strength (31 gms. per liter, 
\ normal), using phenolphthalein suspended in water as indicator (the usual alco- 
holic solution of phenolphthalein will not serve, because the alcohol is at once at- 
tacked) in order to determine the amount of boric acid to be added to the rest 
of the filtrate. (Each cubic centimeter of N/2 boric acid calls for 3 gms. boric 
acid to be added.) An excess of boric acid should be avoided as it favors the 
liberation of hypochlorous acid and renders the solution less stable. It is best 
to add slightly less than the calculated amount. The concentrated solution 
thus prepared contains about 4 per cent of sodium hypochlorite and should be 
mixed with 7 parts of water before use. It can be kept for a month without 
serious decomposition. 



26 A HANDBOOK OF ANTISEPTICS 

solution depends upon the varying composition of the bleach- 
ing powder used and cannot simply be deduced from its 
" available chlorine " content, as is frequently stated. The 
proportion of free lime in the bleaching powder is obviously 
as important as its chlorine strength. With some brands 
of bleaching powder retailed in the United States, the fol- 
lowing proportions have proved useful : 

Two hundred grams of bleaching powder (24-28 per cent 
available chlorine) l is shaken well with five liters of water 
and allowed to stand for an hour or two. In a separate 
vessel dry sodium carbonate (94 grams) and sodium bicar- 
bonate (86 grams) are mixed with five liters of cold water, 
and when dissolved, the solution is added to the bleaching 
powder suspension and the mixture well shaken. The pre- 
cipitate of calcium carbonate is allowed to settle and the 
clear supernatant solution is syphoned off and filtered. The 
solution should contain close to 0.5 per cent sodium hypo- 
chlorite and this should be checked by analysis (p. 41). If 
too strong it should be diluted with water to 0.5 per cent 
strength. 

It is most important to test the solution for free alkali by 
adding a trace of solid phenolphthalein to a little of it. In 
case a red color develops indicating free alkali, the solution 
may still be used if it is previously neutralized either by 
passing carbon dioxide through the solution or by adding a 
little boric acid, until the alkaline reaction is abolished. But 
in making further quantities of the solution, using the same 
sample of bleaching powder, alkalinity may be avoided by 
reducing the quantity of sodium carbonate and correspond- 
ingly increasing the bicarbonate. 

Sodium hypochlorite, whether prepared according to the 
preceding formulae or according to other methods that will 
occur to the chemist, e.g. from salt by electrolysis (p. 116) 
or from liquefied chlorine gas, when used in neutral solution 

1 Or an equivalent amount of stronger bleaching powder. 



ANTISEPTICS OF THE CHLORINE GROUP 27 

at 0.5 per cent concentration is found to be a valuable anti- 
septic for the treatment of infected wounds. Its action is 
extremely rapid and then ceases as soon as all the hypo- 
chlorite is decomposed, hence the methods for using the solu- 
tion efficiently must provide for its frequent renewal. A 
short account of the technique advocated by Carrel and 
Dehelly will be found on p. 13. For further details, reference 
may be made to their book and to the papers noted below. 1 

The hypochlorite solutions possess the valuable property 
of assisting in the rapid dissolution of necrotic tissue, doubt- 
less owing to their ability to react with proteins with the 
formation of soluble products. They possess a slight but 
definite haemostatic action but are actively hemolytic and 
should not be injected intravenously. The hypochlorites 
are extremely reactive substances chemically, and should 
neither be heated above 37 C. or used with other antiseptics 
nor with alcohol nor ether. 

It is rather difficult to give useful figures for the germicidal 
effects of sodium hypochlorite since so much depends on the 
capacity of the medium to decompose the hypochlorite 
before it can complete disinfection. Most pyogenic organ- 
isms suspended in water are killed at a concentration of less 
than 1 : 100,000, while in serum about 1 : 1500 is necessary. 
Blood decomposes the hypochlorites rapidly, so that 1 : 300 or 
more may be necessary before sterilization is complete. The 
action of sodium hypochlorite and eusol on a mixture of pyo- 
genic and other organisms in a blood serum muscle extract 
mixture is recorded on p. 85. The lethal concentration under 
the stated conditions is probably a little less than 1 : 1000. 

m 1 Le traitement des plaies infect es. A. Carrel & G. Dehelly. Masson et 
Cie., Paris, 1917. 

Carrel, Dakin, Daufresne, Dehelly, and Dumas. Presse Medicale, Oct. 11, 
1915. 

Turner, Bull, de l'Acad. de Med., 74, No. 38, 1915. 

Depage, A., Bull, et Mem. Soc. de chir. de Paris, 42, p. 1987, 1916. 

Lyle, H. H. M., Journ. Am. Med. Assoc, Jan. 13, 1917. 



28 . A HANDBOOK OF ANTISEPTICS 

The extraordinary rapidity of its action in concentrations 
even lower than those employed for surgical purposes is well 
illustrated. 

Chloramine-T 

Chloramine-T is the abbreviated name for sodium toluene- 
sulphonchloramide. 1 It is a crystalline, odorless substance 
containing 12.6 per cent of chlorine. It is readily soluble 
in water and the solutions, which have a bitter taste, are 
stable, neither moderate exposure to heat nor light caus- 
ing appreciable decomposition. In equimolecular solutions 
its germicidal activity is about four times that of sodium 
hypochlorite. This may, perhaps, be explained by the fact 
that the chlorine is already linked to nitrogen and is less 
rapidly appropriated through reactions with proteins and 
other substances in the wound secretions. The relatively 
slight solvent action of chloramine-T on necrosed tissue 
supports this view. Its antiseptic efficiency is prolonged 
by this reduction in reactivity. Its germicidal action is 
rapidly exerted and in most respects it resembles the hypo- 
chlorites closely, though decidedly less irritating than the 
latter. An idea of its potency may be gathered frorrl the 
accompanying tables, although in some respects a better 
indication is given by the results recorded in a later chapter 
(p. 86). 

In another series of experiments note was taken of the 
speed of disinfection. Horse blood serum (2 cc.) or 0.7 per 
cent Witte's peptone (2 cc.) was inoculated with staphy- 
lococcus aureus and subsequently treated with weak chlora- 
mine-T solutions (1 cc). The results show clearly that the 
rate of disinfection is very rapid, the maximum effect being 
observed in a few minutes. In those cases where disinfection 
was incomplete and no active antiseptic persisted in the 
mixture, subsequent growth took place (p. 30). 

1 Dakin, Cohen, and Kenyon, Brit. Med. Journ., Jan. 29, 1916. 



ANTISEPTICS OF THE CHLORINE GROUP 



29 



Table showing the Germicidal Action of Chloramine-T on Several 
Common Organisms 

Two drops of a fresh culture of the organisms were suspended in 5 cc. of 
fluid, either water or 50 per cent horse serum, and the antiseptic was allowed to 
act two hours at room temperature. The mixtures were then subcultured. In 
comparison, a few figures for sodium hypochlorite and phenol are added. 





Chloramine-T 


Sodium 
Hypochlorite 


Phenol 


Staphylococci in water . . 


1 : 500,000 — 


1 : 500,000 — 


1 : 250 - 




1 : 1,000,000 + 


1 : 1,000,000 + 


1 : 500 + 


Staphylococci in serum 


1 : 1,500 - 


1 : 1,500 - 


1: 50 - 




1 : 2,500 + 


1 : 2,000 + 


1 : 100 + 


B . pyocyaneus in water 


1 : 200,000 — 


1 : 100,000 — 


1 : 200 — 




1 : 400,000 + 


1 : 1,000,000 + 


1 : 400 + 


B. p3 r ocyaneus in serum . 


1 : 1,250 - 


1 : 2,500 — 


1: 25 - 




1 : 2,000 + 


1 : 5,000 + 


1: 50 + 


Streptococci in water . '. 


1 : 1,000,000 — 






Streptococci in serum . . 


1 : 2,500 — 
1 : 5,000 + 






B. capsulatus in water . . 


1 : 1,000,000 — 






B. capsulatus in serum 


1 : 2,500 — 
1 : 5,000 + 







Complete sterilization is indicated by 
survived. 



-, while + indicates that organisms 



Chloramine-T may be used for wound treatment in solu- 
tion, in the dry state as an impregnation of gauze, or in a 
cream-like paste, all these methods having proved efficient 
and useful. 

In Solution. — At first, solutions containing as much as 
4 per cent of chloramine-T were used in the treatment of 
wounds. Although there were no cases reported to show 
that these were unduly irritating, subsequent experience 
has shown that there is no occasion for exceeding a concen- 
tration of 2 per cent. For, as chloramine-T has, weight for 
weight, as great a germicidal power as sodium hypochlorite, 
a 2 per cent solution is considerably more potent than the 



30 



A HANDBOOK OF ANTISEPTICS 



Medium 


Concentration of 

Chloramene-T in 

the Mixture 


Time of 
Action 


Bacterial 

Count 

( 1 Drop = ^cc.) 


Blood Serum .... 


1 : 1000 





1751 






5 mm. 





Blood Serum .... 


1 : 2000 


15 mm. 
45 







1831 


Blood Serum .... 


1 : 3000 


5 mm. 
15 min. 
45 min. 



5 min. 
15 min. 


9 1 
15 
49 
1509 
96 1 
82 


Peptone 0.7 per cent . . 


1 : 5000 


45 min. 

5 min. 


211 

9360 








15 mm. 





Peptone 0.7 per cent . . 


1 : 6000 


45 mm. 



5 min. 
15 min. 



1323 

13 l 
12 






45 mm. 


14 



solution of sodium hypochlorite usually employed (0.5 per 
cent). The 2 per cent solution may be used for the treat- 
ment of septic wounds, using the same methods as employed 
for the hypochlorites. 

It must be borne in mind that in severe septic conditions 
much of the chloramine-T is promptly decomposed by the 
secretions. This is true of all antiseptics of the chlorine 
group and is even more marked in the case of hypochlorites 
than with chloramine-T. While this circumstance undoubt- 
edly exerts a favorable influence through the destruction of 
substances of a toxic nature, it reduces the germicidal 
activity of the solution. It follows also that where sepsis 
is less marked, much weaker solutions may be employed. 

1 All antiseptic decomposed. No active chlorine present. 



ANTISEPTICS OF THE CHLORIXE GROUP 31 

In the eye, for example, a solution of one part of chloramine-T 
in a thousand parts of normal saline solution will exert a 
satisfactory germicidal action, while i : 500 may prove 
rather irritating to the inflamed conjunctiva. Here, because 
of the constant irrigation by tears, there is chance for only 
a moderate accumulation of septic products likely to reduce 
the strength of the application. In cystitis, the tolerance 
often appears to be less than in the eye, and it is advisable 
to start treatment with a weak solution, increasing the 
strength according to the degree of tolerance manifested. 
In chronic urethral infections, 1 : 500 can be used for 
the initial injections and the concentrations subsequently 
increased. Similar considerations apply to the irrigation 
of the pleural cavity in empyema. The use of chloramine-T 
for the disinfection of meningococcus carriers is referred to 
on p. 100. 

From the foregoing statements it will be evident that the 
choice of strength to be used must be left to the judgment 
of the surgeon. In practice it is advisable to keep a 2 per cent 
solution in stock and to dilute this, if necessary, either with 
water or, in case of considerable dilution, with normal saline 
solution. A 2 per cent solution is slightly hypotonic and 
when an approximately isotonic medium is desired, normal 
saline solution should be used as a diluent. Chloramine-T, 
like hypochlorites, has a corrosive action on most metals 
and should not be used for the sterilization of instruments. 

/;/ Impregnated Gauze. — Chloramine-T is well adapted 
to this use, for which very few substances of high antiseptic 
value have proved successful. It is possible, for example, 
to incorporate as much as 25 per cent of the weight of the 
gauze. This is a much larger amount than is advisable, 
5 per cent being adequate. Obviously the gauze should not 
be moistened before use lest the antiseptic be washed out 
because of its ready solubility. It can be used dry for 
lightly packing and" subsequently moistened if necessary 



32 A HANDBOOK OF ANTISEPTICS 

when in position. Impregnated gauze finds application in 
wounds of recent origin in which the chief object is to pre- 
vent progress of infection. It is particularly adapted to 
use in cases of industrial accident where treatment can be 
promptly instituted. Where frequent renewals of the anti- 
septic or irrigation are called for, it is superfluous. 

In Soap Paste. — None of the chlorine group of anti- 
septics can be used in ointments containing fats or oils as 
these rapidly withdraw the active chlorine with the pro- 
duction of inert compounds. It is possible, however, with 
the exceptionally stable chloramine-T to obtain a prepara- 
tion which can be used as an acceptable substitute for oint- 
ments. A preparation of this sort introduced by Daufresne, 1 
which has been extensively used, contains 0.7 per cent to 1 per 
cent of chloramine-T dissolved in water containing 5-10 per 
cent of sodium stearate. It is important that the mixture 
should not contain any substance which can unite with chlo- 
rine to form an inert compound. For this reason the stearate 
used must be free from any fatty acid of an unsaturated series. 
The paste is not oleagenous. It has a creamy consistency and 
can be spread readily. As the antiseptic is dissolved in the 
water constituting the chief bulk of the paste, it has ready 
access to the parts treated. 

An investigation of the effect of this chloramine-T paste 
in sterilizing moderately infected wounds and maintaining 
asepsis in wounds previously sterilized by other means, has 
been published by Carrel and Hartmann. 2 Its use does not 
delay the rate of cicatrization. 

Preparation. — Chloramine-T was first prepared by Chat- 
taway 3 by the action of sodium hydroxide upon toluene- 
sulphondichloramine (dichloramine-T, vide infra).* A more 

1 Journ. Exper. Med., 26, p. 91, 1917- 2 Ibid., p. 95, I9i7- 

3 Trans. Chem. Soc, 87, p. 153, ioo5- 

4 Chloramine-T is manufactured by Messrs. Boot, Island Street, Nottingham, 
England, and several other firms. It is also marketed by the Abbott Laboratories, 
Chicago, under the name of Chlorazene. 



ANTISEPTICS OF THE CHLORINE GROUP 33 

economical method of preparation consists in dissolving 
toluene-p-sulphonamide (i mol.) in 5 per cent cold alkaline 
solution of sodium hypochlorite (1.2 mol.), warming gently 
if necessary, filtering, and adding 1^ vols, of saturated salt 
solution. The chloramine-T crystallizes out of solution as 
a white glistening meal of crystals and is filtered off, washed 
with salt solution, and dried in the air. The product con- 
tains three molecules of water of crystallization. If it is 
desired to free the substance from adherent salt left from the 
process of preparation, it may be obtained pure from a hot 
concentrated solution, from which on slow cooling it will 
separate in large crystals. The purity of a given sample 
may be determined by titration with decinormal thiosulphate 
solution, as described on p. 42. 

The reaction which takes place in the above preparation 
.may be represented as follows : 

CH 3 CH 3 

+ NaCIO =1] + H 2 



SO2NH2 S0 2 NaNCl 

Toluene-p-sulphonamide Chloramine-T 

DlCHLORAMINE-T 

Dichloramine-T is the abbreviated name for toluene-p- 
sulphondichlor amine. It is a yellowish white crystalline 
substance possessing a sweetish, rather pungent chlorous 
odor. It is stable in the solid state especially when kept in 
the dark. Water dissolves only traces of it, though it is 
readily soluble in most organic solvents except paraffin or 
petroleum. It has an intense germicidal action (pp. 86, 93) 
corresponding to its high content of active chlorine, but it 
is difficult to find perfectly satisfactory solvents for it which 
will yield stable solutions. Up to the present, the best 



34 A HANDBOOK OF ANTISEPTICS 

medium that we have been able to find is a mixture of euca- 
lyptol and paraffin oil, both previously treated as described 
on p. 37 to reduce their avidity for chlorine. A more highly 
chlorinated eucalyptol prepared according to a formula worked 
out by Paul Lewis may prove preferable for many purposes. 

Dichloramine-T was originally used in oil solution for 
nasopharyngeal disinfection (p. 102), but more recently it 
has found a wider application in the treatment of infected 
wounds. The results obtained in the treatment of industrial 
injuries by W. E. Lee x and his colleagues in Philadelphia 
and by J. E. Sweet 2 in war wounds, have been extremely 
satisfactory. 

It will be well, perhaps, to refer first of all to the way in 
which the antiseptic action of dichloramine-T in oil-solution 
is exerted. It is well recognized that antiseptics incorporated 
with or dissolved in oily substances usually possess little if 
any antiseptic activity because intimate contact with the 
infected matter is hindered by the oil. When, however, 
such oil-solutions of dichloramine-T as will be described are 
brought in contact with aqueous media, the partition co- 
efficient between the oil and the water is such that a 
certain amount of the dichloramine-T passes into the water 
and there exerts its germicidal action. The amount of 
dichloramine-T thus passing from the oil is enhanced by the 
presence in the aqueous medium of substances capable of 
taking up chlorine. So that the oil solution serves as a store 
of the antiseptic which is drawn upon to maintain the germi- 
cidal activity of the aqueous medium with which it is in 
contact. Thus the amount of active antiseptic leaving 
the oil solution is, to a considerable extent, dependent upon 
the rate at which it is used up in the aqueous medium. 

As illustrating the influence of varying conditions on the 
passage of the active chlorine from the oil to an aqueous 

1 Journ. Amer. Med. Assoc, July 7, 1917. 

2 Brit. Med. Journ., Aug. 25, 1917. 



AXTISEPTICS OF THE CHLORINE GROUP 35 

medium, the following experiments may be cited. A 6.5 
per cent solution of dichloramine-T, prepared as described 
later, was mixed (a) with an equal volume of saline, (b) with 
muscle extract, (c) with blood serum. The oil was then 
separated completely after 3 hours. The active chlorine 
stated in terms of dichloramine-T in the saline solution was 
1 : 6000, in the filtered muscle extract 1 : 300, and in serum 
which was mostly coagulated 1 : in. It is thus seen that the 
dichloramine-T dissolved in the oil is in a readily available 
form and direct bacteriological tests following its action on 
bacterial suspensions in blood and muscle extract (p. 186) on 
the organisms of the nasopharynx (p. 102) and on war w r ounds 
have clearly shown its germicidal action to be great. 

Dichloramine-T is employed for surgical purposes in a 6.5 
to 10 per cent solution in treated eucalyptol and paraffin 
oil, or in a 20 per cent solution in specially highly chlorinated 
eucalyptol. The preparation of the substance and solvents 
is described on succeeding pages. It is best applied by means 
of an oil spray, an ordinary hard rubber or all glass " atomizer " 
being best, as metal is slowly attacked. It may also be 
poured into wound cavities and it can easily be introduced 
into sinuses by means of a cotton swab dipped in the solution. 
The amount of the solution needed for each treatment is 
extremely small, 1 to 2 cc. being sufficient for most moderate- 
sized wounds, and it need not be renewed more often than 
once in 24 hours. A minimum of dressings is required and 
they do not stick to the granulation tissue. Dichloramine-T, 
like most other antiseptics of the chlorine group, is an active 
lymphagogue when placed on fresh wounds. As granulation 
tissue develops, the lymph discharge decreases and the w r ound 
becomes comparatively dry. It also possesses in marked 
degree the property of aiding in the removal of necrotic 
tissue. The first application of the oil solution causes a 
smarting sensation, which passes aw r ay in a few minutes. 
Wounds treated with the oil fill rapidly with granulation 



36 A HANDBOOK OF ANTISEPTICS 

tissue of healthy color which shows no tendency to exuberant 
growth nor to become sodden. The early reports as to the 
results of the use of dichloramine-T in oil solution for wound 
treatment are decidedly encouraging and the method appears 
to us to be of genuine value and deserving of extended 
use. 1 

Dichloramine-T in oil solution has been tried extensively 
in the early treatment of injuries, not only of the soft parts 
but of tendons, bones, and joints. At the primary dressing 
of these wounds, after the excision of dead tissue and obvious 
foci of infection, the wound is liberally flooded with the 
oil and then closed without drainage. In Lee's experi- 
ence primary union follows in at least 75 per cent of these 
sutured wounds if treated within three hours of their in- 
fliction. If signs of infection should appear, one or more 
stitches are removed and oil is introduced by means of a 
grooved director to the focus of infection once in 24 hours. 
With wounds treated after a longer interval the question of 
their closure becomes a matter of surgical judgment. Di- 
chloramine-T in oil has also been found decidedly useful in the 
treatment of boils and carbuncles with minimal incision, in 
osteomyelitis, non-tuberculous empyema and in the control 
of post-operative wound infection. More recently it has 
been used by Lee for the treatment of burns, and in these 
cases it is advisable to cover the burn with one layer of a 
coarse-meshed gauze previously soaked in paraffin wax. In 
this way the exudate easily comes through the open mesh and 
the dressings do not stick. It is an advantage if such wounds 
can be exposed to the air. The absence of suppuration and 
freedom of drainage in these cases is significant. 

Preparation of Dichloramine-T . — The following details 
are based on the method employed by Chattaway : Bleach- 
ing powder (350 to 400 gms.) of good quality (25 per cent 

1 At the time of writing more than ten thousand cases have been treated with 
dichloramine-T. 



ANTISEPTICS OF THE CHLORINE GROUP 37 

or more "available chlorine ") is shaken with two liters 
of water in a shaker for an hour and then the mixture allowed 
to settle. The supernatant fluid is siphoned off and the 
remainder filtered. Powdered toluene-p-sulphonamide (75 
gms.) is then added to the whole of the hypochlorite solution 
and shaken till dissolved. The solution is filtered if necessary, 
placed in a large separating funnel, and acidified with acetic 
acid (100 cc.) added in portions. About 100 cc. of chloro- 
form are then added to extract the dichloramine precipitated 
by the acid, and the whole well shaken. The chloroform 
layer is tapped off, dried over calcium chloride, filtered and 
allowed to evaporate. The residue is powdered and dried in 
vacuo. It is sufficiently pure for most purposes without 
recrystallization. 

An alternate method of preparation is as follows : Toluene- 
p-sulphonamide (50 gms.), water (500 cc), crystallized 
sodium acetate (100 gms.), and chloroform (100 cc.) are 
placed in a flask which is immersed in cold water and the con- 
tents treated with chlorine gas to saturation. If necessary, 
more chloroform is added to dissolve the dichloramine com- 
pletely. The chloroform is separated, dried, filtered, and 
evaporated as above described. The yield is practically 
theoretical. The product may be tested by titration with 
thiosulphate solution (p. 42). 

Preparation of Chlorinated Eucalyptol for Use as 
a Solvent for Dichloramine-T. — Eucalyptol (U.S. P. 
or Brit. Pharm.), and not eucalyptus oil, must be used. Five 
hundred cc. are treated with 15 gms. potassium or sodium 
chlorate and 50 cc. concentrated hydrochloric acid for 12 hours 
or longer. It is then well washed in a separating funnel, first 
with water and then with a solution of sodium carbonate, 
to remove all traces of hydrochloric acid. After tapping off 
the aqueous layer, 15 gms. dry sodium carbonate are added 
to the oil and the whole allowed to stand for 24 hours. It is 
then filtered, further dried with a little solid calcium chloride, 



38 A HANDBOOK OF ANTISEPTICS 

which must be allowed to act for a considerable time, and is 
then ready for use. 

The degree of chlorination thus achieved does not affect 
the appearance or limpidity of the eucalyptol. A more 
complete chlorination converts the eucalyptol into a much 
heavier oil, which has been used in preparing more concen- 
trated solutions of dichloramine-T for use in the treatment 
of wounds. Its preparation is as follows : Dry chlorine is 
passed into eucalyptol without cooling, allowing the mixture 
to heat up only moderately. The hydrochloric acid formed 
is permitted to escape freely. After prolonged chlorination 
the specific gravity of the mixture rises to about 1.2 and 
very little more chlorine is taken up at room temperatures. 
The oil is washed with sodium carbonate solution, then 
shaken with a concentrated solution of calcium chloride, 
and finally dried over solid calcium chloride and filtered. 
Solutions of dichloramine-T in this oil are stable for many 
weeks. 

Preparation of Chlorinated Paraffin Oil for Use 
in Diluting Eucalyptol Solutions of Dichloramine : T. 
— Paraffin oils derived from different sources vary greatly 
in their capacity for taking up chlorine. By intensive chlo- 
rination it is possible to break up all grades of the oil. The 
following treatment has been found serviceable for a moderate 
chlorination sufficing to protect the dichloramine-T from 
rapid decomposition : To 500 cc. of pharmacopceal liquid par- 
affin add 15 gms. potassium or sodium chlorate and 50 cc. 
concentrated hydrochloric acid. Expose the mixture to 
light, preferably sunlight, for several hours. Transfer to 
a separating funnel and wash successively with water, a solu- 
tion of sodium carbonate, and again water. The water is 
tapped off as completely as possible and a small quantity 
of solid calcium chloride added to the opalescent oiL After 
standing for several hours about 5 gms. of animal charcoal 
are added and the mixture well shaken. On subsequent 



AXTISEPTICS OF THE CHLORINE GROUP 39 

filtering through paper a clear, sometimes slightly yellowish 
oil is obtained, which is ready for use. 

Preparation or the Dichloramine-T Solution. — The 
constituents of the solution are all stable and may be pre- 
served indefinitely, especially in colored bottles, but the 
solution itself is sensitive to light and should be used only 
for a few days. A definite crystalline deposit as distinct 
from a faint opalescence is evidence of decomposition and 
such solutions should be rejected. Careful protection of the 
oil from light will do much to prevent decomposition. In 
preparing the solution it is necessary to dissolve the di- 
chloramine-T in the chlorinated eucalyptol first and to add 
the paraffin oil last. To hasten the rate of solution, the 
dichloramine-T and eucalyptol may be w r armed for a few 
minutes to 6o°. The mixture as used by Sweet on war 
* wounds was made by dissolving 10 grams dichloramine-T 
in 75 cc. of chlorinated eucalyptol and then adding 75 cc. 
of chlorinated paraffin oil. This mixture contains about 
6.5 per cent dichloramine-T. The proportion of paraffin 
oil may be reduced one half if a stronger (10 per cent) solution 
is required. 

Much stronger solutions than this have been used by Lee. 
The more highly chlorinated eucalyptol referred to on p. 38 
was used as sole solvent, no paraffin oil being added. As 
strong as 20 per cent dichloramine-T solutions are used in 
this way with excellent results for primary dressings with- 
out noticeable irritation. 

The Chemical Determination of the Concentration 
of Chlorine Antiseptics 

It is very desirable that the concentration of solutions of 
the various antiseptics of the chlorine group should be sub- 
jected to analytical control. This is particularly true in 
the case of the hypochlorites and other unstable products. 



40 A HANDBOOK OF ANTISEPTICS 

The methods for doing this are extremely simple and the 
slight extra trouble is well repaid by the resulting certainty 
as to the strength of the antiseptic solutions. 

In principle, the same method is used for all the substances 
mentioned in this chapter. A known quantity of the solu- 
tion or substance is taken and an excess of potassium 
or sodium iodide and of acetic acid is added. Iodine is at 
once liberated in amount equivalent to the active chlorin^ 
of the antiseptic, and this iodine is measured by determin- 
ing the amount of a decinormal solution of sodiun 
thiosulphate necessary to react completely w T ith the iodine. 
The following solutions are required : 

Decinormal Sodium Thiosulphate Solution. — This 
is prepared with sufficient accuracy by dissolving 24.8 grams 
of the pure crystals in water and diluting to 1000 cc. The 
solution is moderately stable, especially if protected from 
light, and will serve for two or three months. Precipitation 
of sulphur in the bottle indicates decomposition and when 
this occurs a fresh lot should be made. 

Each cubic centimeter of this solution is equivalent to : 

0.0127 gram Iodine 

0.00354 gram Chlorine 

0.00262 gram Hypochlorous Acid 

0.00372 gram Sodium Hypochlorite 

0.01407 gram Chloramine-T 

0.006 gram Dichloramine-T 

0.00675 gram Halazone 

Potassium Iodide Solution. — A 10 per cent solution in 
water is used. Its exact strength is immaterial and sodium 
iodide may be employed equally well. 

Acetic Acid. — A 10 per cent solution of the pure acid. 

Starch Paste. — Prepared by boiling about 0,1 gram 
starch with 100 cc. of water, cooling, and allowing to sediment. 
The clear solution is poured off and used as an indicator for 



AXTISEPTICS OF THE CHLORINE GROUP 41 

iodine. It must be made fgresh occasionally as moulds are 
apt to grow in the solution. 

Determination of the " Available Chlorine " in 
Bleaching Powder. — A fair average sample from bulk 
is taken and of this 10 grams is exactly weighed out into a 
mortar. The powder is triturated in the mortar with suc- 
cessive small quantities of water and completely transferred 
to a liter flask which is filled to the mark with water. The 
whole is well shaken and allowed to stand for an hour or two. 
Ten cc. of the supernatant liquid is measured with a pipette 
and transferred to a small flask. Five cc. each of the iodide 
and acetic acid solutions are then added. The iodine which 
is now liberated is determined by adding from a burette 
the decinormal sodium thiosulphate solution until almost 
all the iodine has disappeared. A few drops of the starch 
paste are then added and the addition of the thiosulphate 
continued until the blue color just disappears. The 10 cc. 
of bleaching powder solution is equivalent to o.i gram of 
the solid substance and as each cc. of thiosulphate = 0.00354 
gram available chlorine, the percentage of available chlorine 
in the bleaching powder is found by multiplying the number 
of cubic centimeters of thiosulphate used by 0.00354 X 
1000 = 3.54. Thus if a particular sample of bleaching 
powder treated as described required 9.4 cc. of N/10 sodium 
thiosulphate, the available chlorine would be 33.3 per 
cent. 

Titration oe Sodium Hypochlorite Solutions or 
Eusol. — Ten cc. of the solution is treated with 5 cc. each 
of the iodide and acetic acid solutions and then titrated 
with sodium thiosulphate, as described above. Each cubic 
centimeter of thiosulphate used represents 0.00372 gm. 
sodium hypochlorite or 0.00262 gm. hypochlorous acid. The 
quantity of hypochlorite or hypochlorous acid in 100 cc. will 
be given by multiplying the number of cubic centimeters of 
thiosulphate used by 0.0372 or 0.0262 respectively. 



42 A HANDBOOK OF ANTISEPTICS 

Estimation or Chloeamine-T and Dichloramine-T. — 
These substances are examined for their content of active 
chlorine, as described for the hypochlorites with the slight 
difference that in order to facilitate the reaction of the 
chloramines with the iodide, it is well to add a little chloro- 
form (5-10 cc.) or carbon tetrachloride before titrating. The 
method of calculation follows from the fact that each cubic 
centimeter of thiosulphate used in the titration is equivalent 
to 0.01407 gm. crystallized chloramine-T or 0.006 gm. dichlora- 
mine-T. It will be noticed that one molecule of chloramine-T 
liberates two atoms of iodine and dichloramine-T liberates 
four atoms of iodine. The reason for this is that each atom 
of chlorine in the antiseptic is equivalent to a molecule of 
hypochlorous acid, each of which liberates two atoms of iodine 
from an acidified iodide solution. 

HCIO + 2 HI = I 2 + HC1 + H 2 



CHAPTER III 
THE PHENOLIC GROUP OF ANTISEPTICS 

Many of the common antiseptics belong to this group 
and have been long employed for routine disinfection both 
for surgical and hygienic purposes. It cannot be said 
that any new properties of value have been observed in 
this group as the result of their employment in war 
surgery. 

Phenol has found steady advocates ever since Lister 
adopted it as an aid' in obtaining his brilliant successes in 
antiseptic surgery. Early in the present war the mistaken 
notion seemed to prevail that a decision as to the general utility 
of antiseptics could be attained by treating infected wounds 
receiving a minimum of preliminary surgical treatment with 
pure phenol and observing whether infection supervened. 
In the light of present experience it seems hardly necessary 
to say that wound sterilization cannot often be effected by such 
methods. The use of destructive coagulants such as phenol 
in high concentration is undoubtedly undesirable and apt 
to lead to conditions favorable to the growth of anaerobes. 
On the other hand, phenol at a concentration of 2.5 to 5 per 
cent is still in common use and a mixture of equal parts of 
one of these solutions with hydrogen peroxide is viewed 
with favor by many. Phenol and camphor when rubbed 
together in equal proportions give a liquid which is said 
to give useful results in infected cases of long standing. 1 

1 Feldman and Walton, Lancet, Dec. 3, 1916. 
43 



44 A HANDBOOK OF ANTISEPTICS 

The stability of phenol solutions and their clean odor are 
attractive qualities but, while phenol is undoubtedly a good 
disinfectant for many purposes, it does not seem to give as 
good results in the treatment of badly infected wouncs 
as many other antiseptics. The use of alcohol and glycerine l 
as solvents for phenol has not much to recommend it, viewed 
simply as regards disinfection, since both of these solvents 
depress its germicidal activity. Phenol dissolved in vege- 
table oils is almost devoid of germicidal activity as but little 
of the antiseptic leaves the fatty solvent. Much weaker 
solutions dissolved in mineral oil, in which it is sparingly 
soluble, have been found by Lewis and Richards to be 
more effective. The fairly strong inhibitory effect of phenol 
upon phagocytosis has already been referred to, although 
it may be doubted whether this is as important a matter 
as it is sometimes regarded. 

Anthrax spores are remarkably resistant to phenol solu- 
tions and may be viable after four days' immersion in a five 
per cent solution. When acting in the presence of blood 
serum, defibrinated blood, or pus, for reasonable lengths of 
time, e.g. 2 hours, concentrations below 2 per cent are 
relatively ineffective against pyogenic cocci. The rather 
slow but progressive disinfection of a heavily infected mix- 
ture of blood serum and muscle extract, to which one third 
volume of 2 per cent phenol had been added, is well 
illustrated on p. 86. Sterilization was incomplete after 22 
hours although less than 1 per cent of the organism sur- 
vived. The way in which phenol exerts its bactericidal 
action is not understood. Cooper 2 has produced some 
evidence tending to show that chemical reaction between 
the bacterial proteins and phenol is not a sine qua non of 
phenol disinfection and it is suggested that the absorption 
of phenols by bacteria is merely the initial stage in the pro- 

1 Goodrich, Brit. Med. Journ., May 19, 1917. 

2 Biochem. Journal, 7, p. 175, 1913- 



THE PHEXOLIC GROUP OF ANTISEPTICS 45 

cess of disinfection and that the germicidal action which 
follows is due to a de-emulsifying action upon the colloidal 
suspension of some constituent protein essential for the 
stability of the organism. With phenol and its derivatives 
there appears to be an intimate relation between their 
germicidal powers and their protein precipitating capacity. 

Phenol is commonly used as a standard for the measure- 
ment of the efficiency of disinfectants by the Walker-Rideal 
method (page 79). 

Various halogen derivatives of phenol have been proposed 
as antiseptics and while many of them are highly germicidal 
against bacteria suspended in water, they are not particularly 
active in the presence of blood serum or other protein 
material and hence have little to recommend them for wound 
treatment. A great many of these compounds have been 
carefully studied by Bechold and Ehrlich. 1 

Cresols, or methylphenols, occur in three isomeric modi- 
fications. They are more actively germicidal than phenol 
itself and it is stated that a 1 per cent solution of commer- 
cial cresol in water is as active as 3 per cent phenol. 
The commercial mixture goes by the name of " tricresol " 
and is often employed for sterilization of the hands and of 
instruments. Cresol paste, made with lanoline and white 
wax, was recommended by Sir. W. Watson Cheyne 2 and his 
colleagues for the early treatment of infected war wounds, 
but the results obtained early in the present war were gener- 
ally regarded as unfavorable and its use has been discontinued, 

Lysol is prepared by treating the fraction of tar-oils 
chiefly composed of cresols with fat and then saponifying 
with alcoholic soda. Its use for general disinfecting pur- 
poses is well known, but its employment as a dressing for 
wounds is limited, except in veterinary practice. It gives 

1 Zeitschr. F. physiol. chem., 47, p. 173, 1906. 

2 Journ. Royal Naval Medical Service, April, 1915. Lancet, Nov. 21, 
1014, Feb. 27, 1915. 



46 A HANDBOOK OF ANTISEPTICS 

a soapy, frothing solution when mixed with water. Creolin 
is a similar preparation. 

The interesting suggestion has been made by Miss Mary 
Davies 1 that wound infections might be limited to some 
extent by the use of antiseptic substances for impregnating 
the clothing of soldiers. After studying the effect of various 
substances she concludes by recommending the use of a 5 per 
cent solution of "pyxol," a cresol and soft soap preparation 
analogous to lysol. It appears that some bactericidal power 
is retained by cloth so treated after a month's exposure to 
sun and rain and possibly even for a longer period. The 
practical results of these suggestions will be awaited with 
interest. 

Thymol, or propylmethylphenol, has been recommended 
as an antiseptic for surgical purposes but has been prac- 
tically discarded, although dentists find its low solubility an 
advantage in some conditions requiring antiseptic treatment. 
Di-iododithymol is known under the name of aristol. It is 
used to some extent as a substitute for iodoform but is 
unsuited for general use in the treatment of war wounds. 

Salicylic Acid, or orthohydroxybenzoic acid, has received 
much attention as an antiseptic for the treatment of war 
wounds. In particular, a powdered mixture of salicylic 
acid and boric acid, introduced by Sir W. Watson 
Cheyne 2 under the name of borsal, was given an ex- 
tended trial. A mixture of salicylic acid and borax had 
been tried, with ineffective results, as a wound dressing 
by the Japanese in the Russo-Japanese war. Borsal does 
not seem to have achieved much greater success in most 
surgeons' hands, and after fairly extensive trials in France, 
either alone or in conjunction with cresol past, its use was 
abandoned. 3 It appeared that borsal did not effectively 

1 Lancet, Sept. 30, 1916, p. 603. 

2 Brit. Med. Journ., May 22, 1915, p. 912. 

3 Ibid., June 5, 1915, p. 984. 



THE PHENOLIC GROUP OF ANTISEPTICS 47 

check sepsis save in superficial wounds and its use, in the 
opinion of many, was not unattended by danger. It seems 
that the coagulating action of salicylic acid on blood and 
wound exudates impedes free drainage and under these 
conditions, when sterilization has not been complete, the 
possibilities for the growth of anaerobes such as the gas 
bacillus and bacillus of malignant oedema, are considerable. 
The disinfecting action of salicylic acid alone is not great in 
the presence of wound exudates and is, moreover, limited 
by its low solubility, i : 500 in cold water. Addition of borax 
causes a much larger quantity of salicylic acid to dissolve 
owing to the formation of a double salt, sodium borosalicy- 
]ate, which is freely soluble. Solutions of this salt are used 
to a small extent as antiseptic lotions. Alcoholic solutions 
of salicylic acid have been added in small quantity to the 
last funnel full of saline used for irrigating septic wounds. 1 
In this way the salicylic acid is precipitated by the water and 
may be evenly distributed over the surface of the wound. 
This method has been found to be preferable to dusting the 
dry powder on the wound. 

A large number of halogen derivatives of salicylic acid 
have been examined without revealing any particularly 
valuable qualities, although occasionally their germicidal 
propsrties are found to be markedly greater than those of 
salicylic acid. 

/3-Naphthol and Bromonaphthols. — The naphthols 
have antiseptic properties similar to those of the simpler 
phenols but have not been used extensively as wound antisep- 
tics, although /2-naphthol finds some employment as an intes- 
tinal disinfectant and is regarded as valuable in ointments 
for the treatment of skin diseases of parasitic origin. Recently 
/?-naphthol has been used as an antiseptic addition to the 
paraffin wax mixtures used in the treatment of burns. The 
composition of "ambrine," one of the best known of these 

1 L. Garret Anderson and Helen Chambers, Lancet, June 3, 1916. 



43 • A HANDBOOK OF ANTISEPTICS 

mixtures, has not been disclosed by its proprietary owners 
but a product made according to the following formula, due 
to A. J. Hull, 1 is stated to give equally good or better 
results: /3-naphthol 0.25 per cent, eucalyptus oil 2 per cent, 
olive oil 5 per cent, hard paraffin 25 per cent, and soft paraffin 
67.75 P er cent. The mixture may be applied with a broad 
camel hair brush or sprayed on at a temperature of about 
50 C. An illustration of the form of spray used for this 
purpose in the naval service will be found in the British Med- 
ical Journal, August 28, 1917. Resorcinol, 0.25-1 per cent, 
may be used in place of the /3-naphthol. The preparation of 
the mixture is as follows .: Melt the hard paraffin, and add 
the soft paraffin and olive oil. Add the resorcinol dissolved 
in half its weight of absolute alcohol and lastly add the 
eucalyptus oil when the w r ax has cooled to about 55°C. 

Becholdt 2 has carried out experiments with a series of 
bromine derivatives of /?-naphthol and finds that several of 
them have germicidal properties of a high order when tested 
against pyogenic cocci in water suspension. The tribrcm- 
/3-naphthol in particular was found to be especially active, 
killing staphylococci at a dilution of 1 : 250,000. Our own 
experiments have, however, indicated a materially lower 
germicidal value. This substance has been made commer- 
cially and endorsed as an efficient antiseptic agent for the 
treatment of infected wounds. It is, however, not suitable 
for military surgery as it is very sparingly soluble and is 
not significantly more active in the presence of blood serum 
than ordinary /?-naphthol. When dissolved in alcohol and 
tested against staphylococci suspended in serum, the lethal 
concentration is reduced to 1 : 800 or less. 

Picric A cid (trinitrophenol) . — Picric acid is made use of 
more for the treatment of burns than for ordinary infected 

1 Brit. Med. Journ., Jan. 13, igi7. 

2 Zeitschr. f. Hyg. u. Infekt. Krankh., 64, p. 113, 1909. Zeitschr. f. Angew 
Chem., 22, p. 2033, 1909. 



THE PHENOLIC GROUP OF ANTISEPTICS 49 

wounds. The acid is soluble at room temperature in about 
ninety parts of water and dissolves much more readily in 
alcohol or ether. A i per cent or saturated aqueous solu- 
tion is generally employed for surgical purposes. As a 
first dressing for burns it has proved of the greatest value 
and does much to relieve pain and reduce the risk of sub- 
sequent infection. It is employed either in solution or as 
impregnated gauze or wool. 

The use of picric acid in the treatment of extensive wounds 
is probably to be deprecated, not only because other more 
suitable antiseptics are available but also because the sub- 
stance is decidedly toxic. The germicidal properties of 
picric acid tested against staphylococci or B. colt in aqueous 
media are moderately high ; using the Walker-Rideal method 
of testing, it is found to have a " phenol coefficient " variously 
estimated between 4 and 6. 1 But picric acid is an active 
protein precipitant and it is unlikely to be capable of exert- 
ing very much germicidal action in the presence of serum or 
wound exudates. Direct experiments in this point appear 
to be lacking. The coagulating and hardening effect of 
picric acid due to the power of precipitating proteins is prob- 
ably responsible in part for its successful use in the treat- 
ment of burns. 

1 H. L. Tidy, Lancet, Sept. 11, 1915. 



CHAPTER IV 
SALTS OF THE HEAVY METALS AS ANTISEPTICS 

The metallic salts which are used because of their anti- 
septic properties are mainly those of silver, mercury, bismuth, 
and zinc. With one exception, namely, Rutherford Morison's 
bismuth paste, no very extended use of these substances 
has been made in the present war. In the following chapter, 
their mode of action will be referred to first of all, then their 
germicidal effects and, lastly, some of their applications. 

The germicidal activity of many of these metallic salts, 
when acting upon bacteria suspended in pure water, is ex- 
traordinarily high. But this powerful action is enormously 
reduced as soon as the bacteria are placed in other media 
than pure water. Most of the soluble salts of these metals, 
with the exception of some colloidal preparations, suffer 
from the disadvantage that they are precipi table by pro- 
teins or some of the constituents of wound exudates such 
as phosphates, so that their high initial antiseptic potency 
is soon reduced. It also follows from this fact that the 
metallic salts find their most useful applications under 
conditions which do not lead to their rapid precipitation, 
and indeed under such circumstances they are among the 
most valuable disinfectants. 

There is a good deal of evidence pointing to the belief 
that the metallic ions present in aqueous solutions owing to 
electrolytic dissociation are the chief disinfecting agents * and 

1 Dreser Arch. f. Exper. Path. u. Pharm., 32, p. 456, 1893. 
50 



SALTS OF THE HEAVY METALS 51 

that the undissociated salts as such are of minor importance. 
Thus Kronig and Paul found that the disinfecting action of 
equimolecular quantities of mercuric chloride, bromide, and 
cyanide was in proportion to their ionic dissociation in solu- 
tion. Miss Chick has shown that the laws found to govern 
other examples of disinfection are only applicable to the 
action of mercuric chloride if the concentration of Hg" 
ions is used as the basis of calculation rather than the 
total concentration of the salt. The metallic ions are re- 
sponsible for the ordinary chemical reactions of metallic 
salts in aqueous solution and those metallic compounds 
which do not yield the ordinary chemical reactions for the 
metals, such as many protein and other colloidal prepara- 
tions of the metals, possess inferior disinfecting properties. 
An interesting example of the importance of the metallic 
ions in disinfection is shown by the following observation : 
Two silver salts were compared as regards their action on 
staphylococci in water and blood serum. One of these 
salts, silver fluoride, which undergoes dissociation with 
formation of silver ions, killed in two hours staphylo- 
cocci in water at a concentration of less than i : 10,000,000. 
The other salt was the double cyanide of silver and sodium 
which, on solution in water, gives few if any metallic ions 
but remains in solution as a complex aggregate. When 
tested against staphylococci in water, the lethal concentra- 
tion was only about 1 : 5000. Thus it will be seen that silver 
fluoride solution containing metallic ions is, under the con- 
ditions of the experiment, two thousand times as active as 
silver sodium cyanide which gives few or no metallic ions. 
When the same salts were tested against staphylococci 
suspended in blood serum, the lethal concentrations were 
much more closely approximated, being about 1 : 7000 and 
1 : 3000 respectively. It appears probable that the metallic 
ions of the salts exert their disinfecting action by reacting 
chemically with the protein or other constituent of the 



52 A HANDBOOK OF ANTISEPTICS 

bacterial protoplasm, and indeed in some cases this phenom- 
enon has been actually observed. Salts of the heavy metals, 
even when present in a nutrient medium in extremely small 
amount, are capable of exerting an inhibitory influence upon 
the growth of bacteria, so that in all estimations of the germi- 
cidal action of these salts it is essential to avoid carrying 
over any of the antiseptic into subcultures. In addition 
to the inhibitory action upon the growth of bacteria exer- 
cised by traces of metallic salts, Miss Chick 1 has noted another 
phenomenon exhibited by this class of disinfectant. " If 
bacteria are subjected to the action of i : iooo, i : 10,000 or 
even weaker solutions of mercuric chloride, there is an in- 
terval during which some at least of them may be resusci- 
tated by the timely administration of an antidote (in this 
case a sulphide solution), but if this antidotal treatment is 
not employed, no amount of subsequent dilution beyond 
the limits when inhibition occurs, can prevent the death 
of the organism. It would seem that the mercuric salt has 
been already absorbed by the bacteria and possibly formed 
some combination with its substance, not however to a - 
sufficient extent to prevent recovery if a large excess of the 
sulphide solution be employed. - " In one case it was observed 
that in a 24-hour culture of B. paratyphosiis some individuals 
at least were able to manifest vitality after contact with 
5 per cent mercuric chloride solution for four minutes, pro- 
vided ammonium sulphide was promptly applied as antidote. 
There is an extensive literature dealing with the germi- 
cidal action of the metallic salts, but most of the results 
are not comparable with one another owing to variations 
in the technique employed in making the tests. Even slight 
variations in the composition of the medium in which these 
substances act produce enormous variations in the results 
of the experiments. Thus we have found the apparent 
activity of mercuric chloride against staphylococci in a 

1 Journ. of Hygiene, 8, p. 92, 1908. 



SALTS OF THE HEAVY METALS 53 

hard tap water to be only one fifth of that shown when 
distilled water was employed, while with silver salts the 
results are even more liable to variation. The following 
extracts from published experiments are given simply as a 
rough guide to antiseptic potency, but for details, the original 
papers must be consulted. 

The use of mercuric chloride as a disinfectant practically 
dates from Robert Koch's l experiments published in 1881. 
Somewhat later, Geppert showed that Koch had overes- 
timated the antiseptic value of the salt through error in tech- 
nique involving the carrying over of mercury salts into 
subcultures. Geppert's experiments were adversely criti- 
cized by von Behring 2 but have been essentially substan- 
tiated by subsequent workers. The action of mercuric 
chloride upon spores, especially those of B. anthracis, has 
-been most carefully studied by Kronig and Paul, 3 Madson 
and Nyman, 4 and Miss Chick. 5 A chart from Miss Chick's 
paper, in which the figures are taken from Kronig and 
Paul's experiments, gives a good idea of the general character 
of the results, though many more details will be found in 
the original papers. 

As exemplifying the action of mercuric chloride on a vege- 
tative form, B. paratyphosus, the following chart is repro- 
duced from Miss Chick's paper. 

The action of mercuric chloride upon bacteria is not nearly 
so rapid as is commonly thought to be the case. Miss Chick 
found that B. paratyphosus could withstand the action of 
five per cent bichloride for four minutes, and staphylococcus 
aureus, for fifteen minutes, if at once treated with a sulphide 
antidote. 

But these experiments relate only to the action of the 
antiseptic on suspensions of organisms in an aqueous medium. 

1 Ueber Desinfection, Mittheil. Kaiserl. Gesendheitsamt, Vol. I. 

2 Zeitschr. f. Hygiene, g, p. 396, 1890. 3 Ibid., 25, p. 1, 1897. 
4 Ibid., 57, p. 388, 1907. 5 Journ. of Hygiene, 8, p. 92, 1908. 



54 



A HANDBOOK OF ANTISEPTICS 



When working in a blood serum medium, its activity is much 
reduced, while in the presence of whole blood, pus, or muscle 



CHART I 



4000 


















c 






















3500 
























3000 
2500 














































° 1 


s 


















1500 




















No. of bacteria 
o o 




o * 


\ 


i 












I 


rv 






>■— -0-. 


-X 


' -( 


h r 


> ■€ 



30 4.0 50 

Time in minutes 

Illustrating the results of Kronig and Paul's experiment. Continuous curve, 
disinfection of anthrax spores with 2.1 per 1000 HgCb. Dotted curve, 
disinfection of anthrax spores with 1.1 per 1000 HgCla. 

extract much higher concentrations are required for effec- 
tive action. A single new experiment may be quoted 
to illustrate this (see p. 87 ). Mercuric chloride (1 cc. 
1 : 1000) was added to blood serum (1 cc.) and 50 per cent 
muscle extract (1 cc.) previously heavily inoculated with 



SALTS OF THE HEAVY METALS 
CHART II 



55 



1-0 
0-9 






















08 












7 




























































o 
o 
























w 












.2 o 1 

u 

si 










ft 



50 100 150 200 

Time in minutes 



250 



Times taken for disinfection of B. paratyphosus with varying concentrations 
of mercuric chloride, H2S being used as antidote. 



56 A HANDBOOK OF ANTISEPTICS 

staphylococcus aureus. The final concentration of mer- 
curic chloride was therefore i : 3000. The number of sur- 
viving organisms was estimated at varying intervals of time. 1 
The experiment was conducted at 32 C. Although at the 
end of five minutes almost 90 per cent of the organisms 
were killed, the mixture was not sterilized completely at the 
end of three hours. 

Among other salts of mercury that are employed for surgical 
purposes, are mercury potassium iodide, cyanide, oxycyanide 
and double zinc cyanide. "Mercuric biniodide," i.e. mer- 
curic potassium iodide, resembles mercuric chloride closely in 
germicidal properties but is regarded as less irritating. The 
other salts mentioned are less active under most conditions. 

The germicidal action of most silver salts closely resembles 
that of the corresponding mercuric compounds. Silver 
chloride is insoluble and hence ineffective, but silver nitrate 
appears to resemble mercuric chloride fairly closely. Its 
action on B. paratyphosus has been quantitatively studied 
by Miss Chick with results that resemble the curve given 
on p. 55. Silver cyanides, colloidal silver, and various 
organic compounds which yield few silver ions on solution 
in water all appear to have inferior germicidal properties, 
although some of them find useful application in civil prac- 
tice. For their preparation, properties, and uses, reference 
must be made to textbooks of pharmacology. 

Experiments on the action of silver nitrate solution and 
argyrol on considerable quantities of staphylococci and 
other organisms suspended in equal parts of blood serum 
and muscle extract are recorded on p. 87. Silver nitrate 
(1 per cent) added so that the final concentration was 
0.33 per cent killed about 95 per cent of the organisms 
in six hours but failed to sterilize completely in 24 hours. 
Argyrol with a final concentration of 5 per cent did sterilize 

1 Potassium sulphide was added as " antidote " to prevent the carrying over 
of active mercury salts to the subcultures. 



SALTS OF THE HEAVY METALS 57 

in 24 hours but many organisms were still present at the 
end of six hours. 

The salts of bismuth do not appear to have been examined 
very carefully as regards their bactericidal action, although 
their action in checking undesirable fermentations in beer 
worts was discovered long ago. Most normal bismuth salts 
are more or less rapidly decomposed by water with formation 
of insoluble basic salts, and some of these have found effec- 
tive use as mild antiseptics capable of slow but prolonged 
action. 

Zinc salts have long been known to have antiseptic prop- 
erties and the chloride especially has been used by Lister, 
Kocher, and others. Its germicidal action is far inferior 
to that of most mercury or silver salts. A 5 per cent 
solution is ineffective against anthrax spores but a 2.5 per 
cent solution is reported effective against most vegetative 
-forms in a reasonably short time, when acting in an aqueous 
medium. Its activity is however much influenced by the 
medium in which it acts, since it is very readily precipitated 
by proteins, phosphates, etc. An experiment in which 3 
per cent zinc chloride was added to a mixture of equal 
parts of blood serum and muscle extract inoculated with 
staphylococci and other organisms, so that the final con- 
centration of zinc chloride was 1 per cent, showed that about 
one sixth of the organisms survived at the end of an hour 
and a half and that complete sterilization was not quite 
accomplished after 24 hours (p. 88). 

Uses or Metallic Salts in War Surgery. — The prac- 
tical uses of the metallic disinfectants in the treatment 
of infected war wounds has not been extensive with the ex- 
ception of a paste containing bismuth subnitrate which will 
be referred to later. Mercury salts alone have almost no 
advocates, although a certain amount of gauze impregnated 
with Lister's double cyanide of mercury and zinc is still em- 
ployed, with the object of preventing re-infection of the wound 



58 A HANDBOOK OF ANTISEPTICS 

and in the hope of controlling further progress of the existing 
infection. A preparation of mercuric chloride and malachite 
green introduced by Fildes, Rajchman, and Cheatle has, 
however, given useful results, especially in chronic suppurat- 
ing wounds. An account of this preparation will be found 
in the section on dyes (p. 61). 

Silver nitrate at i : iooo was tried for some time, but the 
darkening of the treated wounds on exposure to light was a 
drawback and the results appeared to be only moderately 
good, so that at the present time it is scarcely used at all. 

Zinc salts have a marked caustic and coagulant action 
and, as is well known, are frequently employed on account 
of these effects on torpid ulcerations, fistulous tracts, etc. 
Lister long ago made use of zinc chloride solution in many 
infected conditions and noted the fact that the precipitation 
of zinc compounds in the coagulated surface of the wound 
was generally sufficient to prevent recurrence of sepsis. 
Strong solutions of zinc chloride, up to as high as 10 per cent, 
were believed useful, particularly by Belgian surgeons, when 
used as wet dressings on wounds which had been freely 
incised subsequent to the development of gas gangrene infec- 
tion. This treatment, which is a drastic one, has been largely 
supplanted by irrigation with some form of hypochlorite 
solution. 

By far the most useful metallic salt antiseptic so far em- 
ployed in the present war is the bismuth paste introduced 
by Rutherford Morison. 1 This is made by mixing bismuth 
sub nitrate (i part) and iodoform (2 parts) with sufficient 
liquid paraffin oil to make a thick paste of such consistence 
that it may be readily spread in a thin layer with the 
help of a spatula or spoon. This mixture, which is com- 
monly known as " B. I. P.," was first of all recommended for 
the treatment of infected war wounds in which suppura- 
tion was already established. But it is now used to a con- 
1 Lancet, Aug. 12, 1916, p. 268. 



SALTS OF THE HEAVY METALS 59 

siderable extent for the treatment of fresh wounds, partly 
owing to its ease of application and the fact that frequent 
redressing is usually unnecessary, although adequate pro- 
vision must be made for free drainage. When casualties 
are unusually numerous and speed of treatment becomes 
of great -practical importance, the paste is said to be particu- 
larly useful. 

The method of using the paste for suppurating wounds 
is essentially as follows : After appropriate surgical treat- 
ment, the wound cavity and surrounding skin is carefully 
mopped with alcohol. The wound is then rilled with the 
paste, and dressed with gauze which is covered with an 
absorbent pad, which in turn is held in position with stick- 
ing plaster and a bandage. This dressing requires no change 
for days or weeks if the patient is free from pain and con- 
stitutional disturbances. Should, however, discharge come 
through the dressing, the stained part must be soaked in 
alcohol and a fresh gauze dressing, wet with alcohol, applied 
as a further covering. In order to redress the wound, wool 
soaked in alcohol is used to wipe away the sticky, dirty 
looking discharge. The wound and a small area of skin 
is again plastered with paste and this in turn covered with 
gauze dressing, pad, and bandage, as before. 

It is claimed that the results following the use of this 
paste are notably good in fractures of the long bones, and the 
ease of dressing as well as its infrequency and the absence 
of pain are valuable features. 

Precise bacteriological analysis, of the effects of the paste 
upon infected wounds has not yet been supplied, so that 
final judgment as to the bactericidal value of the mixture 
must be delayed. It must not be forgotten that the con- 
stituents of the mixture are not innocuous and that iodoform 
poisoning, particularly when the drug is placed in closed 
cavities, is not uncommon. However, iodoform poisoning 
seems to be very rarely encountered with the present mixture. 



60 A HANDBOOK OF ANTISEPTICS 

The paste appears to be very slowly absorbed and may cause 
disturbance long after apparent healing of the wound. A 
small sinus may form and iodoform suspended in a brownish 
fluid may escape, but the exit wound heals promptly. In 
the majority of cases, however, it is slowly absorbed and by 
means of X-rays the slow disappearance of B.I. P. in bone 
cavities, etc., can be readily followed. 



CHAPTER V 
DYES AS ANTISEPTICS 

A large number of dyestuffs possess germicidal properties, 
although until recently they have been employed for the 
destruction of blood parasites such as trypanosomes rather 
than bacteria. 

Malachite green, used in conjunction with mercuric chloride, 
was recommended early in 191 5 in a report to the Medical 
Research Committee, by Fildes, Rajchman, and Cheatle, 1 
and has been used fairly extensively, especially in the naval 
service. A 2 per cent solution of malachite green in 80 per 
cent pure alcohol is mixed with an equal volume of a 2 per 
cent solution of mercuric chloride in 80 per cent alcohol. The 
two solutions are best kept apart until needed for use. 
The mixture, as Micklethwaite has shown, contains a double 
compound consisting of one molecule of malachite green and 
two molecules of mercuric chloride. This compound ap- 
pears to be readily dissociated in contact with the tissues. 
The malachite green is reduced by living tissues to the leuco- 
compound and therefore becomes invisible in a wound but 
may still maintain its activity. Sloughs and necrosed tis- 
sue, however, do not reduce the dye. The mixture is gen- 
erally applied by means of a spray and is surprisingly non- 
irritating when the concentration of the mercury salt is 
considered. It has been found particularly useful as a skin- 
disinfectant and for the treatment of superficial wounds, but 
it has also proved of value in cases of osteomyelitis, septic 

1 Lancet, 1915, ii, p. 165. 
61 



62 A HANDBOOK OF ANTISEPTICS 

fractures, and burns. Experiments on the germicidal effect 
of malachite green when acting on organisms either in a 
blood medium or blood serum-muscle extract (p. 89) lead 
us to the conclusion that the value of the dye as a wound 
antiseptic probably has been much overestimated. Media 
such as those noted, to which malachite green had been added 
to a 1 : 1000 concentration, readily underwent putrefaction. 
These conclusions, however, simply refer to the dye itself and 
not to its compound with mercuric chloride as employed by 
Cheatle and his colleagues. 

Malachite green belongs to the group of triphenylmethane 
dyes and may be represented by the following formula: 




(CH 3 ) 2 N< >-C— < >N(CH 3 ) 2 

OH 

It may be prepared by a variety of methods, one of which, 
due to Doebner, 1 consists in heating dimethylaniline (2 
mols.) with zinc chloride and benzotrichloride (1 mol.). 
The zinc salt obtained by this method is commonly con- 
verted into the oxalate which is the usual commercial 
form of malachite green. Browning has shown that some 
oxalates of dyestuffs, including malachite green and brilliant 
green, are more harmful to phagocytosis than other salts, 
so that it might be desirable to employ some soluble salt of 
malachite green other than the oxalate for antiseptic purposes. 
Certain other members of the triphenylmethane group of 
dyes are known to possess definite bactericidal action, as 
shown by Dreyer, Kriegler, and Walker. 2 Hexamethyl violet, 

1 Liebigs Annalen, 217, p. 250. 2 Journ. Path, and Bact., 15, p. 133* I 9 IC - 



DYES AS ANTISEPTICS 63 

also known as " crystal violet," hexaethyl violet, and brilliant 
green are all credited with marked germicidal properties. 
The latter dye has been used to some extent by Browning 
and his surgical colleagues. Our own experiments do not 
indicate that brilliant green is nearly so powerful a germicide 
as claimed by Browning (cp. pp. 90, 95), although it 
doubtless may be of value in the treatment of certain types 
of wounds. A particularly exuberant growth of bright red 
granulation tissue is observed to follow its use. Brilliant 
green or tetraethyldiaminotriphenylcarbinol has a constitu- 
tion similar to that of malachite green but with ethyl 
groups replacing the methyl groups of the latter. It is 
prepared like malachite green by heating diethylaniline with 
zinc chloride and benzotrichloride. 

Acriflavine, Trypaflavine, or Flavine. — This sub- 
stance was first prepared by Benda 1 at Ehrlich's instigation 
in 191 1 and was found to have a marked therapeutic effect 
on trypanosome infections. The systematic name for the 
compound is 3-6 d iamino-10-me thy 1-acridinium chloride and 
is represented by the following formula : 




H 2 N \/\ /\/ NH 2 

/N\ 
CH 3 CI 

The preparation of the compound was protected by patents 
and the registered trade mark " trypaflavine" was assigned to 
it. In order to avoid this name Browning and his colleagues 
used the simple term "flavine/' but as a vegetable yellow dye 
of the same name has long been known this designation was 
1 Ber. deutsch. Chem. Gesell., 45, p. 1787, 191 2, 



64 A HANDBOOK OF ANTISEPTICS 

unfortunate. To avoid these and other important technical 
difficulties the Medical Research Committee has recom- 
mended 1 that so far as Britain is concerned the substance 
should be officially known as "acriflavine" and under this 
name various firms have obtained licenses to manufacture 
it. 2 In this way monopoly from the exploitation of the 
substance under fancy names has been obviated. 

Acriflavine has been claimed by Browning and his associ- 
ates 3 to be a most powerful antiseptic and it has found 
application in the prophylactic treatment of fresh wounds as 
well as in cases where suppuration has developed. It has 
also been employed in the disinfection of the nasopharynx 
of carriers of the meningococcus. Our own experiments on 
the germicidal action of acriflavine lead us to regard it as 
distinctly more active under most conditions than either mala- 
chite green or brilliant green (p. 97), though its rate of 
disinfection is decidedly slow. Direct observations on the 
bacterial count of infected wounds treated with 1 : 1000 
acriflavine confirm our belief that the germicidal action 
of the substance has been overestimated. 

One of the most remarkable properties of acriflavine is 
chat its germicidal action is apparently enhanced by admix- 
ture with serum, though greatly diminished by pus. Relative 
to its bactericidal power, the dye is less detrimental to phago- 
cytosis than most other antiseptics and it has but little in- 
jurious action on the tissues, but on the other hand its 
germicidal action is exerted decidedly more slowly than 
that of some commoner antiseptics. Its solutions may be 
boiled and can even be heated to 120 in the autoclave. 
It is generally used in 1:1000 solution in 0.8 per cent 
salt solution and may be employed* for swabbing or syringing 
septic wounds once or twice daily according to the acuteness 

1 Brit. Med. Journ., June 9, 1917, p. 769. 

2 Acriflavine is, we believe, being manufactured by Messrs. Boot, Island St., 
Nottingham, and doubtless by other firms. 

3 Brit. Med. Journ., Jan. 20, 1917, p. 73. 



DYES AS ANTISEPTICS 65 

of the condition. Gauze soaked in the solution may be 
placed next the wound and a protective covering put over the 
whole to hinder evaporation. Several ounces of i : iooo 
acriflavine solution may safely be left in the tissues or peri- 
toneal cavity. It may also be injected with a serum syringe 
into inflammatory areas. The opinion has been expressed 
that the special uses of acriflavine for particular purposes 
have still to be defined in relation to other antiseptics and 
to the operative methods of surgery, to which it can be at 
best only a valuable aid. 

The following table, taken from Browning's paper, con- 
tains the results of bacteriological tests made with the 
various dyes mentioned. In judging of these results it 
must be borne in mind that relatively very small numbers 
of bacteria were employed (o.i cc. of a i : 20,000 dilution 
in saline of a 24-hour peptone water culture) and that the 
action of the antiseptics was allowed to continue for 24 to 48 
hours before examination. 1 A loopful of the mixture before 
adding the acriflavine is stated to yield twenty or more 
colonies. Such a low concentration of organisms in an old 
wound would indicate approaching surgical sterility, and 
moreover a survival of any number less than 5 per cent 

1 In the original paper by Browning and his colleagues certain antiseptics 
of the chlorine group, e.g. chloramine-T, eusol, chlorine water, and Dakin's 
solution, are included and an attempt is made to determine what is termed " anti- 
septic potency" as expressed by the ratio: 

Lethal concentration in serum of the substance in question 
Lethal concentration in serum of chloramine-T 

These experiments are valueless since the chlorine antiseptics were added first 
to media, either peptone, water, or serum, which in the dilutions employed 
promptly decomposed most of them. For such experiments it is essential to 
add the disinfectant last, as in practical use, and there is little value in continuing 
experiments with rapid-acting unstable chlorine antiseptics for more than a few 
hours. The experiments as described give a false impression of the relative po- 
tency of acriflavine and similar dyes. The original statements by Dakin, Cohen, 
Kenyon, and Daufresne as to the germicidal action of hypochlorites and chlora- 
mine-T in water and serum have been repeatedly confirmed by ourselves and 
many others. 



66 



A HANDBOOK OF ANTISEPTICS 



of the organisms in the test would have a good chance 
of being overlooked. As already stated, our own tests with 
heavily infected mixtures indicate much feebler germicidal 
action than that shown in the fcllowdng table. 



Antiseptic 


Staphylococcus Aureus 
Lethal Concentration 


Bacillus Colt 

Communis, Lethal 

C oncentr ation 


Concentra- 
tion Which 
Inhibits 
Phagocy- 
tosis 


In Peptone 
Water 0.7 % 


In Serum 


In Peptone 
Water 0.7 % 


In Serum 


Malachite 

green 
(oxalate and 

sulphate) 
Brilliant 

green 
(sulphate) 
Crystal 

violet 
Acriflavine 


1 : 10,000,000 

1 : 10,000,000 
1 : 4,000,000 
1 : 20,000 


1 : 40,000 

1 : 30,000 
1 : 400,000 
1 : 200,000 


1 : 20,000 

1 : 130,000 
1 : 8000 
1 : 1300 


1 : 1000 

1 : 35oo 
1 : 8000 
1 : 100,000 


1 : 7000 

1 : 2000 
1 : 7000 
1 : 500 



The preparation of acriflavine on a moderately large scale 
requires a considerable degree of chemical skill. Several 
methods for the synthesis of acriflavine and related acridine 
derivatives 1 are known but only one of them need concern 
us here. The working details must be sought in the 
original communication. 2 

Aniline and formaldehyde unite to give a polymeric sub- 
stance, anhydrof ormaldehy de-aniline (i ) . When heated with 
aniline hydrochloride it undergoes a curious rearrangement 
with the formation of p-diaminophenylme thane (2). This 
substance is nitrated with nitric and sulphuric acids so as to 
give a dinitro derivative (3) ; which in turn is reduced with 

1 Bucherer's Chemie der Tierfarbstoffe, 19 14, may be consulted for informa- 
tion concerning the synthesis of acridine derivatives. 

2 Benda, Ber. deutsch. Chem. Gesel., 45, p. 1787, 1912; D. R. Patents, Kl. 
22b, 230412 and 243085, Casella and Co., 1911-1912. 



DYES AS ANTISEPTICS 



67 



tin and hydrochloric acid. On heating the reaction mixture 
in an autoclave to 135 , 3-6-diamino acridine is obtained (4). 
In order to obtain the methyl derivative the latter compound 
is first converted into its di-acetyl compound to protect 
the amino-groups and then treated in nitrobenzene solution 
at 1 7 5 with the methyl ester of toluene sulphonic acid. 
The product is then hydrolyzed, when acriflavine (hydrochlo- 
ride) = 3-6-diamino-io-methylacridine (5) results: 



■H 2 N 




►H 2 N ^ jNNk, >NH 2 ->H 2 N 
2 2 

(3) 




NH 2 



->H 2 N 




NH 2 



Apparently the germicidal properties of acriflavine are 
shared by a number of other acridine derivatives and one of 
these, diaminoacridine, (4) is easier and cheaper to manufac- 
ture and appears to be equally desirable. It will be known 
as " proflavine." A detailed publication on this substance 
is expected shortly. Experiments on its germicidal action 
are included in Chapter VII. 



CHAPTER VI 
MISCELLANEOUS ANTISEPTICS 

In this section brief reference will be made to a few anti- 
septics which cannot be included in preceding sections. It 
will be impossible to treat these substances systematically, 
and instead brief notes as to the chief characteristics of the 
substances is all that will be attempted. 

Hydrogen Peroxide and other Peroxides. — Hydro- 
gen peroxide is not held in very high repute as a germicide 
but it has certain other qualities which render it decidedly 
valuable. When tested against relatively small quantities 
of staphylococcus aureus or B. pyocyaneus in water, a 
concentration of 0.03 per cent actual H 2 2 , i.e. a 1 per 
cent dilution of the pharmacopoeal product, may suffice to 
sterilize in two hours, while in blood serum about double this 
concentration may be necessary. But these conditions are 
entirely artificial and unrelated to what happens when 
hydrogen peroxide is applied to a septic wound. Blood, 
pus, and muscle juice contain an enzyme "catalase" which 
rapidly brings about the decomposition of hydrogen peroxide 
with liberation of gaseous oxygen. This rapid decomposition 
with evolution of gas soon decomposes all the peroxide and 
its disinfecting action comes to a speedy end. The mechanical 
effect of the disengagement of gas is often a valuable prop- 
erty and is made use of in loosening sticky secretions, 
washing away pus, or loosening adherent dressings. For 
such purpose it is of great value but it is important that 
its transient germicidal effect should be recognized. 

68 



MISCELLANEOUS ANTISEPTICS 69 

A great variety of other peroxides have been put forward 
as disinfectants, but most of them are of doubtful practical 
value. Two of the most active of these substances are the 
benzoyl hydrogen peroxide and benzoyl acetyl peroxide dis- 
covered by Baeyer and studied by Freer and Novy. The 
latter compound has been used to some extent but the un- 
stable character of both the substance and its solutions 
has prevented its extensive employment. Dibenzoyl per- 
oxide has been recommended as an antiseptic but it is a 
sparingly soluble, practically indifferent compound of no 
significant germicidal value. 

Ozone. — This substance has recently been used by 
Stoker l for the treatment of infected wounds, especially 
cavities and sinuses in bone injuries. An Andreoli ozonizer 
is used to produce the ozonized air which is allowed to act 
for about fifteen minutes or such shorter period as pro- 
duces a superficial glazing of the wound surfaces. De- 
tails of ozone concentration and bacteriological controls 
are not yet available, although the results are stated to 
be good and the formation of excessive granulation tissue is 
avoided. 

Iodine. — Extensive use is made of iodine as a germicide- 
Its action is powerful and prompt when the conditions are 
such that the antiseptic has free excess to the microorganisms. 
Ampoules containing tincture of iodine have been used 
largely in the present war as a first aid treatment to be 
applied by the wounded soldier. In general the results as 
regards the prevention of sepsis have been disappointing, 
mainly it would appear because the injured man is not 
apt in most cases to put the iodine where it can reach the 
focus of infection. The presence of much blood and pro- 
longed oozing are of course inimical to the exercise of 
germicidal action. Usually little more is accomplished 
than a fair cleansing of the adjacent skin. 

lancet, Oct. 21, p. 712, 1916. 



70 A HANDBOOK OF ANTISEPTICS 

The use of iodine as a skin disinfectant introduced by 
Stretton x in 1909 is widely practiced and undoubtedly it has 
great value for this purpose. A 2 . 5 per cent solution is usually 
strong enough and alcohol is generally employed as the 
solvent. Seventy per cent alcohol is preferable to stronger 
spirit and it is important to use pure alcohol as otherwise 
iodoacetone and other products are apt to be formed, which 
are very irritating to the eyes of the operator. 

Light petroleum and heavy mineral oils are also used as 
solvents for iodine and act well. They have the advantage 
of being stable and cheap. Dichlorethylene (i.e. acetylene 
dichloride) has also been proposed but is much more ex- 
pensive. 

The use of iodine for the antiseptic treatment of large 
war wounds is now practiced much less frequently than 
before, although it was given a thorough trial in the early 
days of the war. It has been found much too irritating for 
repeated application and not infrequently objectionable after 
effects such as severe neuritis have been observed. Its 
strong coagulating action on proteins is also an objection- 
able feature. In general it may be said that iodine will be 
found most useful when the conditions are such that rapid 
and complete sterilization may be effected by a single appli- 
cation as in skin disinfection or small surface wounds. 

An experiment illustrating the rapid effect of 2 per cent 
iodine solution on staphylococci and other organisms sus- 
pended in a blood serum-muscle extract medium will be 
found on p. 86. A 1 per cent solution added to an equal 
volume of blood heavily infected with streptococci did not 
kill all the organisms in one hour according to Emery. 
Against staphylococci suspended in water, about ten million 
per cubic centimeter, 1 : 100,000 iodine is effective in two 
hours, while in blood serum 1 : 1000 is required, according 
to our own observations. 

1 Brit. Med. Journ., Aug. 14, 1909, May 22, 1915. 



MISCELLANEOUS ANTISEPTICS 71 

Borates, Perborates, and Boric Acid. — These sub- 
stances while possessing almost negligible germicidal prop- 
erties find extensive use when a bland, mildly antiseptic 
lotion is required. Sodium monoborate is a rather strongly 
alkaline salt but is said to be non-irritating to wounds; 
borax, or sodium biborate is less strongly alkaline, while 
boric acid is feebly acid. All of these substances are used 
for restraining the growth of putrefactive organisms rather 
than for disinfection in the true sense. Sodium perborate is 
prepared by adding hydrogen peroxide to borax solutions 
and is stable when preserved in the dry state. It is often 
used as a convenient substitute for hydrogen peroxide. 

Persulphates. — The use of potassium or sodium persul- 
phate has been advocated but actually their disinfecting action 
on pyogenic organisms is feeble. A solution of the sodium 
-salt has been regarded by some as useful for stimulating the 
rate of cicatrization of wounds, though this action can hardly 
be regarded as satisfactorily demonstrated. 

Acids. — Almost all acids possess some germicidal action 
and generally speaking their activity is proportional to their 
" strength." Just as metallic " ions " appear to be the actual 
disinfecting agent when metallic salts are employed, so in 
the case of most acids it is the hydrogen ions which seem to 
be effective. Extensive experiments on this subject have 
been carried out by Bial x and by Winslow and Lockridge. 2 
A few experiments on the action of various acid substances 
on B. typhosus will be found on p. 114. 

Occasionally acids have been employed in wound treatment 
and it has been thought that irrigation with weak lactic acid 
was useful in inhibiting the growth of the gas bacillus in 
infected wounds, but the treatment has not met with general 
favor. 

1 Archiv. exper. Path. a. Pharm., 38, p. 1, 1897. Zeitschr. f. physiol. Chem., 
40, p. 513, 1902. 

2 Journ. of Infectious Diseases, 3, p. 547, 1906. 



72 



A HANDBOOK OF ANTISEPTICS 



Alcohol and Ether. — Both of these substances are 
employed in the treatment of infected wounds but they are 
used as much for the help they afford in the mechanical clean- 
ing of dirty wounds as for any direct germicidal action. 
Most vegetative forms of bacteria may be killed fairly 
readily by 50 per cent alcohol but alcohol of much lower 
or higher strength is less effective, while most spores are un- 
affected by alcohol of any strength. The following table 
contains the results of experiments by Minervini x to deter- 
mine the time required to kill various organisms in alcohol 
of varying concentration. Koch's " thread " method of 
testing the viability of the organisms was used and the 
results are at least comparable among themselves. 



Organism 



Staphylococcus aureus 
B. pyocyaneus . . . 
M. prodigiosus . . 

B. coli 

B. subtilis (spores) 
B. anthracis (spores) 



Dilution of Alcohol 



25% 



12-24 hours 

under i hour 

1 hour 

24 hours 



So % 70 % 



10 mm. 10 mm. living after 
10 min. 10 min. 6 hours 
10 min. 10 min. 6 hours 
i hour i hour living after 
all living after 8 days 
all living after 50 days 



80% 



99% 



3 days 
12 hours 
1 2-24 hours 
24 hours 



It is well known that alcohol or glycerol materially reduces 
the germicidal efficiency of some antiseptics, particularly 
those of the phenol class. Kronig and Paul found that 
phenol dissolved in 98 per cent alcohol was devoid of bac- 
tericidal action when tested against spores. Cooper 2 has 
correlated this fact with a diminished protein precipitating 
action of alcoholic phenol when contrasted with aqueous 
phenol, but whatever the explanation may be it is clear that 
alcohol is not a desirable solvent for phenolic disinfectants. 

1 Cp., Rideal, Disinfection and Disinfectants, p. 322. 

2 Biochem. Journ., 7, p. 175, 1913. 



MISCELLANEOUS ANTISEPTICS 73 

The germicidal properties of ether have recently been ex- 
amined by Topley, 1 who finds that the vapors possess a 
slight but definite action. An exposure to ether vapors of 
one to forty-eight hours was necessary to sterilize agar slants 
on which pyogenic organisms were growing. Liquid ether 
was irregular in its action and good contact with the organ- 
isms was difficult to secure. It is. clear, however, from 
these and from clinical results that any beneficial effects 
following the use of ether in the treatment of septic infec- 
tions is not due to direct disinfection. 2 

Formaldehyde. — The physical and chemical properties 
of formaldehyde, which is obtainable in commerce as a 40 
per cent solution known as " formalin, " make it valuable 
for various forms of disinfection, such as the fumigation of 
rooms, etc. But so far as the treatment of septic wounds is 
'concerned it has not proved particularly successful and is 
now scarcely used for such purposes. It is employed to 
some extent for the sterilization of the hands and instru- 
ments but it is less popular than formerly. A 0.5 per cent 
solution is generally employed for such purposes. It is not 
a very rapid acting disinfectant and when tested in the or- 
dinary way under the standard conditions it has a phenol 
coefficient of about 0.4, but if the time of action is prolonged 
its activity may be somewhat greater than that of phenol. 

Hexamethylenetetramine. — This substance, obtained 
by the action of ammonia on formaldehyde, has but little di- 
rect germicidal action. In acid solution it may be decom- 
posed with liberation of formaldehyde which can thus exert its 
antiseptic action. A large number of derivatives of hexa- 
methylenetetramine have been prepared by Jacobs and 
Heidelberger and many of these are more powerful than 
the parent substance, but their useful application has still 
to be defined. Hexamethylenetetramine itself is not suit- 

1 Brit. Med. Journ., Feb. 6, 191 5. 

2 Cp. Distaso and Bowen, Brit. Med. Journ., Feb. 24, 1917. 



74 A HANDBOOK OF ANTISEPTICS 

able for wound treatment since under these conditions its 
germicidal action is quite inadequate. 

Iodoform. — The use of iodoform in combination with 
bismuth subnitrate and paraffin oil in the mixture known as 
"B. I. P." has already been referred to on p. 58. The sub- 
stance was once considered to be a powerful antiseptic but 
this is no longer believed to be the case. Microorganisms 
may flourish in contact with iodoform but, on the other hand, 
it is not disproved that in contact with living tissues iodoform 
may be slowly decomposed with the formation of products 
of genuine antiseptic value. Apart from the paste referred 
to, its use in surgery is diminishing, probably on account of 
occasional unpleasant poisoning effects that may follow its 
free use, especially in confined cavities. Many odorless sub- 
stitutes for iodoform are known under various trade names, 
but in general their action appears to be similar to that of 
iodoform itself. 

Permanganates. — The potassium salt is principally used 
for irrigation, 1 : 1000, in gonorrhoea. It is an active germi- 
cide under conditions not involving rapid decomposition , 
by excess of organic matter. All the permanganates are 
strong oxidizing agents and as soon as they are reduced to 
manganese salts their disinfecting action ceases, so that 
their maximum germicidal effects are transitory. They are 
unsuitable for septic wound treatment on account of their 
rapid reduction but have many other useful applications. 

Quinine. — The hydrochloride of this alkaloid has been 
recommended by Kenneth Taylor l as a dressing for sep- 
tic wounds, especially those infected with the B. cero genes 
capsulatus. The substance is used in 0.1 per cent aqueous 
solution with the addition of 0.1 per cent hydrochloric acid 
or 1 per cent alcohol to inhibit precipitation of the base. 
Tested in vitro Taylor finds that it is about ten times as ef- 
fective as phenol against the gas bacillus although, on the 

1 Lancet, Sept. 4, 1915. Brit. Med. Journ., Dec. 25, 1915. 



MISCELLANEOUS ANTISEPTICS 75 

other hand, it is used on wounds in much lower concentra- 
tion than is usual with phenol. Serum or pus is stated to 
have no very marked action in reducing the germicidal 
activity of quinine. The solution is used either as a wet 
dressing or for continuous drip instillation. A curious fact 
noted by Taylor in connection with the use of quinine is 
that while there appears to be a decrease in B. cero genes 
capsulatus infection there was a progressive increase in the 
appearance of B. pyocyaneus, and laboratory tests showed 
that the latter organism was the most resistant to quinine 
of the common pyogenic bacteria. 

An endeavor has been made to utilize the combined an- 
aesthetic and antiseptic properties of mixtures of quinine and 
urea for wound treatment, but this has failed to secure much 
favor. 

Chinosol. — This compound was originally considered to 
be potassium oxyquinoline sulphonate or a double salt of 
this with potassium sulphate, but is now stated to be 
neutral oxyquinoline sulphate. It possesses a strong inhib- 
itory action on the growth of many microorganisms and 
failure to recognize this fact fully led to exaggerated claims 
as to its germicidal potency. A full report upon this sub- 
stance has been made by the Council on Pharmacy and 
Chemistry of the American Medical Association. 1 The 
general conclusions arrived at are essentially as follows : As 
regards staphylococcus, aureus and B. typhosus chinosol is 
more strongly antiseptic than phenol and about equal to 
mercuric chloride, but as a germicide in watery solution it is 
somewhat inferior to phenol and vastly inferior to mercuric 
chloride. In acid broth the findings were still less favorable 
to chinosol. 

These conclusions make it doubtful if much direct germi- 
cidal effect on wounds can follow its application, though it 
is possible that the substance might find useful employment 

1 Cp. Journ. ; Am. Med. Assoc, May 28, 1910. 



76 A HANDBOOK OF ANTISEPTICS 

as a wound dressing with the object of restraining the 
growth of organisms and preventing reinfection. It does 
not appear to have been used extensively in the present 
war. 

Acetanilide and its Derivatives. — This substance has 
often been recommended as an antiseptic, 1 but our own ex- 
periments have showm its germicidal properties when tested 
against moderate quantities of staphylococci to be so low that 
it can hardly exert any effective disinfection in wounds. A 
half per cent solution failed to sterilize staphylococci in water 
in two hours, while in blood serum its antiseptic action is 
negligible. Recently E. F. Greene 2 has revived the ques- 
tion of its possible use in infected war wounds, applied as a 
dry powder, but this can hardly be endorsed, not only on 
account of its poor antiseptic action but also because symp- 
toms of poisoning have been stated to follow its use. Its 
free application to extensive wounds would certainly not be 
without danger to many individuals who are fairly susceptible 
to this drug. 

Various derivatives of acetanilide in which the hydrogen 
of the benzene ring has been replaced by chlorine, bromine, 
or iodine have been advocated as antiseptics from time to 
time. Parabromacetanilide is sometimes known under the 
name of " asepsin " or " antisepsin " and is stated to have 
anodyne properties. Our own experience indicates that 
none of these derivatives are sufficiently active disinfectants 
to be of value in wound treatment. 

1 Cp. Beck, New York, Med. Journ., March 19, 1893. 

2 Brit. Med. Journ., May 29, 1915, p. 928. 



CHAPTER VII 
METHODS OF TESTING ANTISEPTICS 

The testing of substances for their antiseptic and germicidal 
power is fraught with innumerable pitfalls. It is possible 
to take almost any substance and by carefully choosing the 
conditions under which it acts, make it appear to possess 
germicidal potency, and conversely, it is equally possible to 
take valuable germicides and by observing their action 
'under unreasonable conditions, make them appear inert. 
The necessity for choosing methods of testing in some de- 
gree in conformity with the mode of use of the substance 
and under conditions not too remote from those under which 
it is proposed to employ it, is of fundamental importance. 

The simplest conditions are presented in the determination 
of the lethal concentration of a germicide acting upon micro- 
organisms suspended in water. The chief variables in such 
a determination are the number of organisms taken for 
each experiment, and the temperature and time of action. 
With regard to the number of organisms, it is desirable to 
use a fairly large quantity, partly because spontaneous death 
of the bacteria then becomes less of a factor and also because 
the test becomes a more rigid one. In general, some such 
concentration as a hundred million bacteria to the cubic 
centimeter will be found appropriate. The effect of tem- 
perature on the rate of disinfection has already been con- 
sidered, also the speed of action will be found to vary 
enormously with different substances and with varying 
media, 

77 



78 A HANDBOOK OF ANTISEPTICS 

One of the oldest methods used for the determination of 
lethal concentrations is the so-called "thread" method 
devised by Koch. Sterile raw silk thread cut into portions 
about a centimeter long are soaked in a broth culture or 
aqueous suspension of the required organisms and then dried. 
These threads are soaked for an arbitrary time in varying 
concentrations of the antiseptic, then removed and trans- 
ferred to sterile broth, which on incubation will show whether 
or not the organisms were killed. One of the main objections 
to this method is the mechanical carrying over of adhering 
antiseptic into the broth medium, so that inhibition of growth 
frequently occurs even though the organisms are not killed, 
and the substance appears to be a more effective germicide 
than is actually the case. Failure of the antiseptic to 
promptly penetrate to all parts of the thread may lead to 
erroneous estimates of its germicidal power. 

A later method, which is often called the "garnet" method, 
was worked out by Kronig and Paul. Garnets of equal size 
are dipped in an emulsion of the organism — usually spore- 
bearing anthrax bacilli — and then carefully dried so that 
a thin film of organisms is spread over their surface. The 
garnets are then immersed in a solution of the antiseptic, 
which after a definite time is gently washed away and the 
garnets, if need be, treated with an antidote such as am- 
monium sulphide when mercury disinfectants are employed. 
The garnets are then shaken with water to detach a fairly 
constant proportion of the organisms and an aliquot part of 
the fluid is plated in solid media for counting. An alternative 
method using the surface of nutrient agar instead of garnets 
or thread, is described by Bechold and Ehrlich. 1 

But neither the " thread " nor " garnet " method is often 
used now and the simpler method is usually employed of 
mixing known but varying quantities of the disinfectant with 
a constant concentration of organisms and subculturing from 

1 Zeit. physiol. Chem., 47, p. 177, 1906. 



METHODS OF TESTING ANTISEPTICS 79 

the mixture into broth or agar at known intervals of time, in 
order to determine whether disinfection has been completed. 
An application of this method, which is of great value in 
standardizing and comparing different substances with 
regard to their practical uses as disinfectants in hygienic 
work, is that devised by Walker and Rideal : in 1 903 . By this 
procedure it is possible to express the value of a disinfectant 
in terms of a numerical ratio using pure phenol as a standard. 
The original method is substantially as follows : a definite 
amount (usually 5 drops) of a 24-hour broth culture of the 
organism selected, e.g. B. typhosus or B. coli, is added to 5 
cc. of solutions of varying concentration of the disinfectant. 
Subcultures are taken at intervals of 2^ minutes up to 15 
minutes to determine the point of complete sterility. A 
similar series of tests is made with a standard phenol solu- 
tion of such strength that sterilization of the organisms will 
be effected within the time limits of 2\ to 15 minutes. The 
necessary concentration of phenol will usually be about 0.8 
per cent with most cultures of B. typhosus. By comparing 
the concentration of the disinfectant which just effects 
sterilization in a given time with the phenol concentration 
which also effects sterilization in the same time, an estimate 
is formed of the germicidal potency of the substance under 
investigation. Thus if a concentration of 1 : 600 of a par- 
ticular substance is as effective under the above conditions 
and in the same time as 1 : 120 phenol, the " phenol coefficient" 

will be or 5. It is an essential and important fea- 

120 

ture of this method that time of action should be constant 
while the concentration of the disinfectant is varied. The 
reverse conditions, namely, fixed concentration and variable 
time, may lead to entirely erroneous inferences as to ger- 
micidal potency. 

1 Journ. Roy. Soc. Inst. 24, p. 424, 1903. Later details concerning the Walker- 
Rideal method will be found in The Amer. Journ. Pub. Health, Vol. 3, No. 6. 



80 A HANDBOOK OF ANTISEPTICS 

The determination of the " phenol coefficient " of a substance 
furnishes most valuable information, but chiefly as regards 
the relative value of disinfectants to be used in fairly homo- 
geneous aqueous media. Chick and Martin * and many 
others cited in the paper by these authors have sought to 
obtain a more practical estimate of the germicidal value of 
disinfectants by adding organic matter of various kinds 
to the bacterial suspension. For some types of general 
disinfectants the addition of a definite quantity of 
dried feces is recommended, for example, when testing 
those substances destined to be used for the disinfection 
of stools. 

The problem of selecting conditions for studying the action 
of substances to be used as wound disinfectants is much 
more difficult. Useful information is obtained by determin- 
ing the lethal concentration of substances acting on organ- 
isms suspended in blood serum or in blood, or by studying 
their action on pus. Certain precautions must be taken 
in working with these media. With all of them, it is of course 
essential that, as in practical use, the antiseptic solution 
should be added to the mixtures last, for in many cases if 
this is not done totally erroneous results will be obtained. 
It has happened with extraordinary frequency that small 
quantities of labile chlorine antiseptics have been added to 
organic media and then at unstated intervals after all or 
most of the antiseptic has been destroyed, the mixture is 
infected with organisms which naturally grow unchecked. 
Under such conditions, what is really being observed is less 
the germicidal action of the antiseptic than the rate of 
chemical reaction which has taken place between the unstable 
antiseptic and organic media. The results of such a pro- 
cedure are naturally particularly misleading when high 
dilutions of antiseptic are employed in experiments to de- 
termine the lethal concentration. 

1 Journ. Hygiene, 8, p. 654, 1908. 



METHODS OF TESTING ANTISEPTICS 81 

A great advantage in the use of serum is the fact that it 
presents a homogeneous medium of fairly constant com- 
position ; but it is important to bear in mind that many or- 
ganisms, e.g. B. paratyphosu's, are very susceptible to the ac- 
tion of blood serum even though previously heated to 58- 
6o° for an hour. To avoid this complication, liberal quanti- 
ties of an organism such as staphylococcus aureus, which is 
fairly resistant to serum, should be used. When substances 
of high germicidal power are examined for the determination 
of the lethal concentration, and hence low concentrations are 
employed, it will usually be found sufficient to carry out the 
tests in a medium containing 50 per cent or even less of 
serum, since its mass, relative to that of the antiseptic, will 
be very large. 

The testing of the germicidal effects of substances acting 
on organisms suspended in blood would seem the rational 
procedure for examining compounds which might be used 
intravenously. A blood medium has, however, been sug- 
gested as a standard method of testing ordinary antiseptics 
by Emery. 1 The use of a blood medium as a standard is 
particularly adverse to the hypochlorite antiseptics which 
rapidly react with haemoglobin so that a relatively con- 
siderable amount of the former must be added before any 
active antiseptic will persist in the mixture. For these 
tests use may be made of reconstituted blood obtained by 
mixing serum with cells separated from citrated blood, al- 
though for most purposes defibrinated blood or citrated 
blood itself would probably serve. The technique em- 
ployed by Emery is substantially the following : Nine parts of 
" reconstituted blood' ' is mixed with one part of a strep- 
tococcus culture containing about 250,000,000 organisms per 
cubic centimeter. One volume of the infected blood is mixed 
on a slide, using a marked capillary pipette, with an equal 
volume of the antiseptic solution. The mixture is then drawn 
1 Lancet, April 15, 1916, p. 817. 



82 



A HANDBOOK OF ANTISEPTICS 



into the pipette, which is sealed and incubated and subse- 
quently examined to determine whether the contents are 
sterile or whether viable organisms persist. It should be 
noted that apparently the concentrations of antiseptic quoted 
in Emery's table refer to the strength of the solution added, 
so that their actual concentration in the final mixture is half 
of that given. 



Antiseptic 


15 Minutes 


60 Minutes 




Did not kill 


Killed 


Did not kill 


Killed 


Phenol 

Eusol (HCIO) 

Sodium Hypochlorite .... 

Mercuric Chloride 

Mercury Biniodide .... 

Iodine 

Lysol 

Malachite Green 


1 : 70 
[1 : 400?] 
[1 : 200?] 
1 : 100 
1 : 60 
1 : 100 
1 : 40 
1 : 250 


1 : 60 

1: 80 
1 : 40 

? 

1: 30 
1 : 200 


1 : 60 

1 : 100 
1 : 60 
1 : 100 
1 : 150 
1 : 250 


1:50 

1:80 
1 : 40 

? 
1 : 120 
1 :2oo 



The results recorded by Emery do not make the method as de- 
scribed by him appear particularly accurate, since with only 
one exception as high a concentration was necessary to steri- 
lize the mixture in 60 min. as in 15 min., although the act 
of disinfection is a progressive time reaction. Moreover, a 
higher concentration of phenol is recorded as necessary to 
sterilize in sixty minutes than is stated to be effective in 
fifteen. But the use of blood as a medium for studying the 
germicidal action of substances in the presence of cellular 
elements, is of distinct value since, unlike pus, its compo- 
sition is fairly uniform. The blood should be heavily in- 
fected with organisms so that its natural bactericidal proper- 
ties will not effett the end result materially. On pp. 93, 95 
a number of experiments with antiseptics of the chlorine 
group and dyestuffs, acting on staphylococci in blood media, 
adre recorded. 



METHODS OF TESTING ANTISEPTICS 83 

The determination of the disinfecting action of substances 
upon pus is obviously important as bearing on the treatment 
of suppurating wounds. As already stated, the disinfection of 
pus is difficult of accomplishment and relatively high concen- 
trations of antiseptic are necessary. But great difficulties 
are encountered in laboratory experiments on the disinfection 
of pus owing to the enormous variations in its physical con- 
dition and the number of organisms it contains and whether 
these organisms are mainly free or ingested in the leucocytes. 
Rous and Jones x have shown that living phagocytes are able 
to protect ingested organisms from the action of substances 
such as potassium cyanide in the surrounding fluid and even 
from the action of a strong homologous antiserum, and they 
obtained evidence that this protection by phagocytes was 
largely conditioned on their being alive. But even when 
all the leucocytes are dead, there are considerable mechanical 
difficulties in securing good contact with the antiseptic 
solution. An interesting series of experiments on the steri- 
lization of pus by various antiseptics is recorded in a paper 
by Parry Morgan, 2 and some of the results have already been 
cited on p. 8. 

Thus far, we have considered almost exclusively the ques- 
tion of the lethal concentration of disinfectants acting under 
different conditions, but it is becoming constantly more 
evident that much more than this is needed in judging of 
the utility of antiseptics. As already stated on p. 6, the 
act of disinfection resembles in many respects an ordinary 
chemical reaction in which the two reacting components are 
represented by disinfectant and bacterial protoplasm. Now 
the rate of disinfection varies enormously with different 
germicides, and is of course influenced by the relative mass 
of bacteria and disinfectant, as well as by temperature and 
contact between the reacting substances, f The instability 

1 Journ., Exp. Med., 23, p. 601, 1916. i 

2 B. M. J., May 13, 1916, p. 684. J 



84 A HANDBOOK OF ANTISEPTICS 

of many antiseptics, leading to a reduction or disappearance 
of the mass of active antiseptic, must also be reckoned with. 
A knowledge of the speed with which a disinfectant acts is 
essential to an understanding of the conditions under which 
it may be appropriately used. For example, the extremely 
rapid acting aqueous hypochlorites are admirably adapted 
for intermittent instillation in large quantities into wound 
cavities, while when the same solutions are applied as wet 
dressings which are seldom renewed, their action is over as 
soon as the active chlorine has disappeared, and this may be 
a matter of only seconds or minutes. For prolonged action 
when intermittent instillation cannot be practiced, a slower 
acting but more stable antiseptic is likely to give better re- 
sults, or they may be attained by using an oil solution of 
dichloramine-T (p. 34) from which the active antiseptic 
slowly passes from the oil to the aqueous medium on the 
surface of the wound. 

The only satisfactory way to follow the speed of disin- 
fection is to determine the progressive change in the number 
of bacteria in a suitable mixture after varying lengths of 
time. With this end in view, we have made a number 
of time experiments with various antiseptics, using them 
in most cases at about the concentration which is recom- 
mended for wound treatment. The mixtures contained 1 
cc. of horse blood serum, 1 cc. of muscle extract obtained 
by soaking fresh veal with an equal weight of saline and 
then straining through cloth but not filtering, and 2 drops 
of a staphylococcus aureus emulsion obtained by shaking a 
24-hour agar slant of abundant growth with 8 cc. of saline. 
The muscle extract doubtless contained organisms of other 
kinds in fair quantity. After determining the total number 
of organisms present in the mixture by plating an aliquot 
part, 1 cc. of antiseptic solution was added and samples with- 
drawn from time to time and the surviving bacteria esti- 
mated. In the case of chlorine antiseptics, their further 



METHODS OF TESTING ANTISEPTICS 



85 



action after sampling was checked by sodium thiosulphate, 
while potassium sulphide was used as an antidote with the 
salts of the heavy metals. No antidote was used with the 
dyes. The use of the blood serum-muscle extract medium 
was chosen as bearing some similarity to the composition of 
wound exudate and it has the further advantage of being 
easily reproduced in fairly uniform quality. For many 
antiseptics, the addition of muscle extract makes the tests 
much more severe than when blood serum alone is used. 
The experiments were all carried out at 32-35 C. and the 
results are expressed as the number of bacteria present in 
one standard drop of the mixture — 1/40 cc. 

I. Chlorine. Group of Antiseptics 
(Blood Serum-Muscle Extract Medium ; Staphylococcus aureus.) 



Antiseptic Used 


Concentration 


Time of 
Action 


Bacterial 

Count 

1 Drop=2 1 oCC. 


As Added 


In Mixture 


I. Sodium Hypochlorite . 


o.5% 


0.17% 




2 min. 

5 min. 


1 966 OOO 
405 
oi 


II. do. do. . . 


o.5% 
(0.2 cc.) 


0.05% • 
0.09% 




5 min. 
15 min. 
45 min. 

1.5 hr. 
3 hr. 

6 hr. 
24 hr. 


1 966 OOO 
311 200 

157 400 

98 170 
16 120 

I 651 

1587 

1 294 OOO 2 


III. Eusol 


0.27% 




2 min. 
5 min. 
15 min. 


2 I50 OOO 

496 

2 
O 1 



1 Active chlorine present. 



2 Active chlorine absent. 



A HANDBOOK OF ANTISEPTICS 



I. Chlorine Group of Antiseptics — Continued 
(Blood Serum-Muscle Extract Medium; Staphylococcus aureus.) 



Antiseptic Used 


Concentration 


Time of 
Action 


Bacterial 

Count 

1 Drop= 5 1 cc. 


As Added 


In Mixture 


IV. Eusol 


0.27% 
(0.2 cc.) 


0.025% 




5 min. 
15 min. 
45 min. 

1.5 hr. 
3 hr. 

6 hr. 
24 hr. 


2 129 000 

1 310 000 
507 900 
606 200 
641 400 
868300 
983 000 

2 310 000 2 


V. Chloramine-T . . . 


2% 


0.67% 




5 min. 


1 365 000 
o 1 


VI. do 


0.5% 


0.17% 




5 min. 
15 min. 
45 min. 

1.5 hr. 
3 hr. 

6 hr. 


726 300 
998 
191 
7 
4 
3 
o 2 


VII. Dichloramine-T in Oil 
(v, p. 39) 


2% 


0.67% 3 




0.5 min. 


2 020 000 



VIII. do 


2% 


0.67% 4 




5 min. 
15 min. 
45 min. 


1 157 000 

458 700 

294 900 




IX. Iodine in Potassium 
Iodide 


2% 


0.67% 




5 min. 


1 463 000 




The results shown in Table I in which members of the 
chlorine group of antiseptics and iodine are considered, il- 



1 Active chlorine present. 
3 Mixed with platinum wire. 



2 Active chlorine absent. 
4 Not mixed, left to diffuse. 



METHODS OF TESTING ANTISEPTICS 



87 



lustrate their extraordinary speed and completeness of dis- 
infection when added in adequate amount to the infected 
mixture. In Experiments I, III, V, VII, and IX, -J volume 
of the various antiseptic solutions was added at a concen- 
tration no higher and in some cases lower than that com- 
monly employed in wound treatment and in every case 
practical sterility was obtained in less than five minutes. 
When much smaller amounts of hypochlorites were added, 
as in Experiments II and IV, their rapid action is seen to 
cease as soon as all the active antiseptic is decomposed and 
subsequent growth can then take place. In Experiment 
VI, in which weak (0.5 per cent) chloramine-T was used, its 
action is shown to be distinctly more effective than an ap- 
proximately equivalent amount of chlorine in the form of 
sodium hypochlorite or hypochlorous acid (Experiments II 
and IV). 

II. Metallic Salts, Phenol, Hydrogen Peroxide 
(Blood Serum-Muscle Extract Medium ; Staphylococcus aureus.) 





Concentration 






Antiseptic Used 




Time op 
Action 


Bacterial 

Count 

1 Drop= 5 1 oCC. 


As Added 


In Mixture 


I. Mercuric Chloride . . 


0.1% 


0.033% 




5 min. 
15 min. 

45 min. 

1.5 hr. 

3 hr. 
24 hr. 


1 894 000 

254 400 

44920 

12 900 

7258 

2985 




II. Silver Nitrate- . . . 


1% 


o.33% 




5 min. 
15 min. 
45 min. 

1.5 hr. 
3 hr. 

6 hr. 
24 hr. 


786 400 
720 900 
651 700 
530 600 
425 900 
175 600 
38 270 
2 643 



A HANDBOOK OF ANTISEPTICS 



II. Metallic Salts, Phenol, Hydrogen Peroxide — Continued 
(Blood Serum-Muscle Extract Medium; Staphylococcus aureus.) 



Antiseptic Used 


Concentration 


Time of 
Action 


Bacterial 

Count 

1 Drop= 3 1 cc 


As Added 


In Mixture 


III. Argyrol 


15% 


5% 




5 min. 
15 min. 
45 min. 

1.5 hr. 
3 hr. 

6 hr. 
24 hr. 


917 500 

753 500 

655 300 

622 500 

327 600 

9 792 

4693 




IV. Zinc Chloride . . . 


3% 


1% 




5 min. 
15 min. 
45 min. 

1.5 hr. 
3 hr. 

6 hr. 
24 hr. 


1 223 OOO 
868 300 
819 200 

49i 5oo 

211 900 

5i 58o 

3667 

704 


V. Hydrogen Peroxide 


2.89% 


0.96% 




5 min. 
15 min. 
45 min. 

1.5 hr. 
3 hr. 

6 hr. 


1 201 000 
589 800 
819 200 

778 200 

802 800 

1 087 OOO 

I 136 OOO 


VI. Phenol 


2% 


0.67% 




5 min. 
15 min. 
45 min. 

1.5 hr. 

3 hr. 
24 hr. 


1 409 OOO 

1 471 OOO 

819 200 

192 700 

34900 

13 700 

2985 



In the second series of experiments a similar infected 
medium of blood serum and muscle extract was mixed with 
solutions of various metallic salts, phenol, and hydrogen 



METHODS OF TESTING ANTISEPTICS 



89 



peroxide. Mercuric and zinc chlorides and silver nitrate 
produced immediate precipitates. The action of mercuric 
chloride o.i per cent is seen to be more rapid than that of 
phenol 2 per cent, silver nitrate i per cent, or zinc chloride 3 
per cent. Argyrol was tried in very high concentration and 
while acting slowly sterilized completely in 24 hours. Neither 
phenol, silver nitrate, nor zinc chloride sterilized completely 
in 24 hours, although the number of viable organisms was 
less than 1 per cent of those originally present in the mix- 
ture. The effect of hydrogen peroxide is interesting as 
showing an extremely rapid but transitory action which 
reached its maximum in a few minutes, after which the 
organisms grew unchecked. 

III. Dyes 

(Blood Serum-Muscle Extract Medium ; Staphylo coccus aureus.) 





Antiseptic Used 


Concentration 


Time of 
Action 


Bacterial 

Count 

1 Drop= 5 1 oCC. 




As Added 


In Mixture 


I. 


Malachite Green . . 
(Griibler) 


0.3% 


0.1% 




5 min. 
15 min. 

1. 5 hr. 

3 hr. 

6 hr. 
24 hr. 
48 hr. 


821 600 

182 700 

60 920 

8 920 

5 574 

78 200 

1 015 000 

3 706 000 


II. 


do. . . do. . . . 


0.1% 


0.033% 




5 min. 
15 min. 
45 min. 

3 hr. 

6 hr. 
24 hr. 
48 hr. 


2 097 coo 

1 359 coo 

369 100 

211 500 

272 300 

557 coo 

1 096 000 

6 553 000 



90 



A HANDBOOK OF ANTISEPTICS 



III. Dyes — Continued 
(Blood Serum-Muscle Extract Medium; Staphylococcus aureus.) 



Antiseptic Used 


Concentration 


Time of 
Action 


Bacterial 
Count 






As Added 


In Mixture 


iDrop = 3 1 oCC. 


III. Brilliant Green . . . 


o.3% 


0.1% 





810 200 


(Griibler) 






5 min. 
15 min. 
45 min. 

1.5 hr. 
3 hr. 

6 hr. 
24 hr. 
48 hr. 


29 950 

27 410 

20 160 

14 310 

8871 

1 946 

256 600 

658 900 


IV. Acriflavine .... 


0.3% 


0.1% 




5 min. 
15 min. 
45 min. 

1.5 hr. 

6 hr. 
24 hr. 


589 800 

358 400 

249 300 

179 200 

78 210 

704 




V. do 


0.1% 


0.033% 




5 min. 
15 min. 
45 min. 

1.5 hr. 

6 hr. 
24 hr. 


557 100 
218 600 
121 900 
100 800 
81 540 

474 



VI. do 


0.03% 


0.01% 





755 400 
524300 






5 min. 








15 mm. 


319 300 








45 min. 


308 200 








1.5 hr. 


106 500 








3 hr. 


94 840 








6 hr. 


22 780 








24 hr. 


1 113 OOO 


VII. Proflavine 


0.3% 


0.1% 




5 min. 
15 min. 
45 min. 

1.5 hr. 
3 hr. 

6 hr. 
24 hr. 


888 100 

794 500 

744 500 

308 200 

104 800 

66 560 

53 250 

1 242 



METHODS OF TESTING ANTISEPTICS 



91 



III. Dyes — Continued 
(Blood Serum-Muscle Extract Medium ; Staphylococcus aureus.) 



Antiseptic Used 


Concentration 


Time of 
Action 


Bacterial 

Count 

1 Drop =3^00. 


As Added 


In Mixture 


VIII. Proflavine .... 


0.1% 


0.033% 




5 min. 
15 min. 
45 min. 

1.5 hr. 
3 hr. 

6 hr. 
24 hr. 


888 100 
744 500 
655 400 
326 100 
169 900 
154 800 
84 900 
4 


IX. do 


0.03% 


0.01% 




5 min. 
15 min. 
45 min. 

1.5 hr. 
3 hr. 

6 hr. 
24 hr. 


855 400 
761 800 
744 5oo 
469 500 
304 500 
196 400 
93 750 
183 600 



In the third series of experiments the action of malachite 
green, brilliant green, and acriflavine acting in a similar mix- 
ture of blood serum and muscle extract were examined. The 
germicidal action of these dyes, in view of claims made for 
them, proved disappointing. Brilliant green has been recom- 
mended for use by Browning and his colleagues at a concen- 
tration of 0.1 per cent but addition of one third volume of 
brilliant green or malachite green at this concentration gave 
but poor evidence of much germicidal action. Subsequent 
experiments with these two dyes were made at a higher 
concentration, 0.3 per cent. In no case was sterility 
reached and after some hours the organisms grew unchecked . 
It was noted that similar mixtures of blood serum and muscle 



92 A HANDBOOK OF ANTISEPTICS 

extract mixed with these dyes so that the final concentration 
was o.i per cent, on exposure to air readily underwent 
thorough putrefaction. It would appear that under the con- 
ditions of these experiments the dyes in question are readily 
inactivated and that the presence of apparently unchanged 
coloring matter in the mixture is no evidence of the continua- 
tion of any antiseptic action. Similar experiments with 
acriflavine, o.i per cent, showed a slow initial effect but anti- 
septic action was more sustained, and while sterility was not 
attained in six hours, after 24 hours disinfection was com- 
plete. When the concentration of the acriflavine solution 
added was 0.03 per cent more organisms were present at the 
end of 24 hours than at the commencement of the experi- 
ment. The results with proflavine were not widely different 
from acriflavine, although the higher concentrations did 
not effect perfect sterilization in 24 hours. The effect of 
these dyes appears more pronounced when acting in blood 
or blood serum than when muscle extract is present. 

We have also made some experiments in the progressive 
change in the bacterial count when various chlorine antiseptics 
and dyes were added to defibrinated blood. With the excep- 
tion of the sodium hypochlorite and eusol tests these experi- 
ments were made by adding one volume of antiseptic to two 
volumes of freshly drawn sterile rabbit blood heavily inocu- 
lated with staphylococcus aureus. The general conditions of 
the experiments were identical with those preceding. 



METHODS OF TESTIXG ANTISEPTICS 



93 



IV. Chlorine Group of Antiseptics 
(Defibrinated Blood Medium ; Staphylococcus aureus.) 



Antiseptic Used 


CONXENTRATION 


Time of 
Action 


Bacterial 

Count 

1 Drop= 4 1 u cc. 


As Added 


In Mixture 


I. Sodium Hypochlorite . 


o.5% 


o.33% 




5 min. 
15 min. 

45 min. 


573 400 
563 

282 
2 432 


II. do. do. . -. . . 


C5% 


0.3% 




5 min. 
15 min. 
45 min. 


573 400 

1 774 

1485 

19 656 


III. do. do 


0.5% 


0.25% 



45 min. 


232 oco 
89 600 


IV. Eusol ...... 


0.27% 


0.18% 




5 min. 
15 min. 
45 min. 


578 600 

1 562 

2 330 

36 600 


V. do 


0.27% 


0.16% 




5 min. 
15 min. 
45 min. 


8ll OOO 
36 280 
48 250 
51 580 


VT. do 


0.27% 


0.13^ 




5 min. 


260 80O 
143 300 


VTI. Chloramine-T . . . 


2% 


0.67% 




5 min. 
15 min. 


224 50O 

125 

O 


VII. do 


•1% 


o.5% 



45 min. 


235 OOO 
O 


VIII. do 


0-75% 


0.25% 




5 min. 
15 min. 
45 min. 

3 hr. 


I 178 OOO 
20 l6o 

5 451 

4 032 

806 


IX. Dichloramine-T in Oil 


2% 0.67% » 




2 min. 


59 900 
O 



Mixture stirred with platinum wire. 



94 A HANDBOOK OF ANTISEPTICS 

The results of the action of 0.5 per cent sodium hypochlorite 
and of eusol on staphylococci in a blood medium show that a 
relatively large addition of these must be made before any 
marked germicidal effect is produced. Even when two 
volumes of the antiseptic were added to one of infected blood, 
complete sterilization was not quite accomplished, although 
considerably over 99 per cent of the organisms were 
killed. As already stated, the reason for the apparent 
low germicidal action of hypochlorites in a blood medium is 
due to their ready decomposition by the haemoglobin and 
other substances in the blood. From a practical standpoint 
this is not of much importance when methods of intermittent 
instillation of hypochlorite solution into wounds are practiced 
but it serves to emphasize the necessity for the frequent 
renewal of the solution. 

Chloramine-T and dichloramine-T give materially better 
results than the hypochlorites when acting on organisms 
in a blood medium. A comparison of Experiments I and IX 
shows that a concentration of 0.25 per cent of chloramine-T 
in the mixture was as effective as 0.33 per cent sodium hy- 
pochlorite, although the active chlorine in the latter was 
about five times as much as that in the chloramine-T. The 
reason for this difference is to be sought largely in the slower 
rate of reaction between chloramine-T and haemoglobin and 
other proteins, compared with the hypochlorite solutions. 



METHODS OF TESTING ANTISEPTICS 



95 



V. Dyes 

(Defibrinated Blood Medium ; Staphylococcus aureus.) 



Antiseptic Used 


Concentration 


Time of 

Action 


Bactertal 

Count 

1 Drop=3 1 oCC 


As Added 


In Mixture 


I. Malachite Green . . 
(Grubler) 


0.3% 


0.1% 




5 min. 
15 min. 
45 min. 

1.5 hr. 
3 hr. 

6 hr. 
24 hr. 
72 hr. 


1 754 OOO 
107 
88 
66 
82 
35 
36 
13 



II. do 


0.1% 


0.033% 




5 min. 
15 min. 
45 min. 

1.5 hr. 
3 hr. 

6 hr. 
24 hr. 


1 693 OOO 

1 310 oco 

819 200 

208 OOO 

146 900 

86 010 

89 600 

1 219 OOO 


III. Brilliant Green . . . 
(Grubler) 


0.3% 


0.1% 




5 min. 


1 966 OOO 



IV. do. do 


0.1% 


0.033% 




5 min. 
15 min. 
45 min. 

1.5 hr. 
3 hr. 

6 hr. 
24 hr. 


1 630 OOO 

1 261 

563 

755 
717 
857 

2 592 
1 171 OOO 


V. Acriflavine .... 


0.3% 


0.1% 




5 min. 
15 min. 
45 min. 

1.5 hr. 
3 hr. 

6 hr. 
24 hr. 


654 600 

358 400 

108 800 

98 180 

I 472 

203 

17 





96 



A HANDBOOK OF ANTISEPTICS 



V. Dyes — Continued 
(Defibrinated Blood Medium ; Staphylococcus aureus.) 



Antiseptic Used 


Concentration 


Time of 
Action 


Bacterial 
Count 








As Added 


In Mixture 


1 Drop= ¥ \)CC. 


VI. Acriflavine .... 


0.1% 


0.03% 




5 min. 
15 min. 
45 min. 

1.5 hr. 
3 hr. 

6 hr. 
24 hr. 


654 700 

411 900 

225 800 

138 100 

30 640 

1 152 

32 




VII. do 


0.03% 


0.01% 




5 min. 
15 min. 
45 min. 

1.5 hr. 
3 hr. 

6 hr. 
24 hr. 


900 600 
819 200 
508 900 
190 OOO 

83 860 

32 260 

768 

3 


VIII. Proflavine .... 


0.3% 


0.1% 




5 min. 
15 min. 
45 min. 

1.5 hr. 
3 hr. 

6 hr. 
24 hr. 


819 200 

4710 

1638 

474 

304 

113 

12 




IX. do 


0.1% 


0.033% 




5 min. 
15 min. 
45 min. 

1.5 hr. 
3 hr. 

6 hr. 
24 hr. 


819 200 
201 300 
208 OOO 
122 5OO 

53 220 

29 670 

2 995 

2 


X. do 


0.03% 


0.01% 




5 min. 
15 min. 
45 min. 

1.5 hr. 
3 hr. 

6 hr. 


819 200 
753 70c 
852 OOO 

591 400 

394 200 

448 OOO 

48380 








24 hr. 


282 



METHODS OF TESTING ANTISEPTICS 97 

The action of malachite green and brilliant green in 
blood heavily infected with staphylococcus aureus was dis- 
appointing. In only one case, namely with brilliant green, 
did a concentration of o.i per cent in the mixture bring about 
apparent sterility. It is by no means certain that even in 
this case all the organisms were killed, since it was not pos- 
sible to avoid carrying over of some of the dye to the agar 
subcultures, and in the concentration referred to we showed 
that reproduction of living staphylococci actually was in- 
hibited, although they were not killed. It is significant that 
with the exception of the two experiments in which brilliant 
green and malachite green were added in 0.33 per cent solu- 
tion, an initial fall in the number of bacteria was followed 
by practically unrestricted growth. These results are in 
marked contrast to those of Browning and his colleagues 
referred to in Chapter V. The action of acriflavine and 
proflavine in blood, while slow, is seen to be distinctly 
superior to that of the other dyes. 

In judging of the suitability of substances for wound 
treatment, there are of course other factors to be considered 
besides germicidal efficiency. Prominent among these is 
the determination of the concentration at which irritation 
of the skin and other tissues becomes noticeable, also the 
effect of germicidal substances upon phagocytosis and their 
influence if any on the rate of dissolution of necrotic tissue. 
All of these factors may best be investigated in vivo. Ref- 
erence may be made to Colonel Bond's ingenious experiments 
noted on p. 11 with regard to phagocytic activity as influenced 
by antiseptics. 

Laboratory experiments serve primarily for the sorting 
out of substances which are likely to be of value for wound 
antisepsis, and they are particularly important in indicating 
the mode of use for the selected substance which is most 
likely to give successful clinical results, but the final decision 
of the utility of any germicide will necessarily follow from a 



98 A H AN D BOOK OF ANTISEPTICS 

study of its effects upon actual wounds. The treatment of 
comparable infected wounds with various antiseptics, accom- 
panied by daily bacteriological estimations of the degree of 
infection gives most useful results. This method has been 
practiced extensively in the present war and gives valuable 
indications as to the earliest date at which wounds may be 
safely closed by suture or otherwise. The influence of 
antiseptics on the rate of cicatrization of wounds may also 
be followed with the aid of a formula worked out by Du 
Xotiy. 1 The technique of these experimental methods is 
beyond the scope of this book and reference should be made 
to original sources. 

1 Journ., Exper. Med., 24, pp. 451, 461, 1916; 25, p. 721, 1917. 



CHAPTER VIII 

CERTAIN SPECIAL APPLICATIONS OF ANTISEPTICS 

I. The Disinfection of Carriers 

The problem of destroying pathogenic organisms in the 
nasal and upper air passages by direct disinfection is a diffi- 
cult one. There are, undoubtedly, many carriers with 
anatomical abnormalities of the nose, pharyngeal vault and 
tonsils precluding immediate contact with solutions used 
either as sprays or gargles. In such cases there is little pros- 
pect of any antiseptic proving effective. Those organisms 
which are exposed to direct contact with the antiseptic 
solution, may be destroyed, but in the course of time those 
which have preserved their vitality in protected situations 
regenerate the original condition and only a temporary, 
although to that extent beneficial, result is attained. This 
difficulty is encountered in chronic diphtheria carriers, in 
whom deep tonsilar crypts are frequently encountered form- 
ing inaccessible regions in which the bacilli are afforded pro- 
tection. The pneumococcus, existing in the pulmonary air 
passages, is, in those parts, wholly beyond reach. In con- 
sidering the applicability of treatment to individual cases. 
these inevitable limitations must be taken into account. 

In spite of these limitations, chiefly imposed by anatomical 
conditions, valuable results have been obtained, notably 
by Gordon and Flack, with the use of antiseptics in the 
treatment of meningococcus carriers among soldiers. By 
a suitable choice of antiseptic, properly applied, there is no 
doubt that much can be accomplished in limiting the risks 



100 A HANDBOOK OF ANTISEPTICS 

connected with carriers of pathogenic organisms, both to 
themselves and those coming in contact with them. 

The methods that have been employed with most success 
are the following : 

(i) Chamber disinfection using a steam spray and a 
solution of chloramine-T or zinc sulphate (Gordon and 
Flack). Hypochlorites have also been used for chamber 
disinfection, but they have not been found as satisfactory. 1 

(2) Local application of various antiseptics including 
iodine, with either menthol, guaiacol or glycerine, axgyrol 
and hydrogen peroxide, zinc salts, acrinavine, chloramine-T 
and especially dichloramine-T in oil solution. Ferric chloride 
and potassium permanganate solutions have given only mod- 
erate results while formalin appears to be distinctly harmful. 

Gordon and Flack found that many of the antiseptic 
solutions employed as gargles and in hand sprays at the 
time when they commenced their experiments gave un- 
satisfactory results when used on heavily infected chronic 
carriers. This appeared to be due as much to lack of per- 
fect contact as to germicidal inefficiency. They therefore 
employed an inhalation chamber with a steam pressure 
spray or atomizer capable of converting the antiseptic 
solution into a fine mist. The chamber was about 750 to 
1000 cubic feet capacity, and the apparatus was capable of 
spraying a liter of antiseptic solution in the course of tw T enty 
minutes. A working diagram of two forms of these sprays 
will be found in Gordon and Flack's original paper, 2 in which 
there will also be found details of the successful treatment of 
numbers of chronic meningococcus carriers. The prepared 
antiseptic solution used for spraying by Gordon and 
Flack was either chloramine-T, 0.5 per cent, or zinc sulphate, 
1.2 per cent. The chloramine-T gave the better results, 
especially with persistent carriers, but is somewhat less well 

1 Kuster, Deutsch. Med. Woch, 41, p. 11 16, 1915. 

2 Brit. Med. Journ., p. 673, Nov. 18, 1916. 



THE DISIXFECTIOX OF CARRIERS 



101 



tolerated than the zinc salt. The inhalation was carried 
out once daily and on each occasion the carriers were in the 
inhaling room from fifteen to twenty minutes during which 
time they vigorously inhaled the spray-laden atmosphere 
through the nostrils. 

Using a different apparatus with a high pressure air spray,, 
the authors l have examined the antiseptic action of chlor- 
amine-T solution on the mixed bacterial flora of the normal 
nasopharynx. The results are recorded in the following 
table in which, for comparison, the effect of spraying with 
salt solution is included. The figures, which are the aver- 
age of six similar experiments, indicate the total number of 
organisms obtained from plating a swab taken under approxi- 
mately constant conditions and in which any antiseptic 
adhering to the swab was immediately destroyed with 
sterile sodium thiosulphate solution. 

Aqueous Chloramine-T and Neutral Salt Solution 





Number of Colonies Derived from Sample 




First Period 


Second Period 


Time of Treatment 


5 min. 


10 min. 


15 min. 


5 min. 


10 min. 


15 min. 


0.5% chloramine-T 
(control, 43.240) 

Normal salt solution 
(control, 56.738) 


12,352 


707 
121,620 


■ 526 
13,587 


439 
2,137 


153 
10,055 


17 
34,339 



From these and other results, it appeared that exposure 
to the chloramine-T spray for less than fifteen minutes was 
not very effective and that frequent repetition of the treat- 
ment was desirable. Repetition several times daily of the 

1 Brit. Med. Journ., June 23, 1917. 



102 



A HANDBOOK OF ANTISEPTICS 



chloramine-T spray for 15 or 20 minute periods is rather a 
severe procedure, so that as an alternative we have em- 
ployed an oily solution of the related dichloramine-T (see 
p. 33), which has the great advantage that its action is much 
more prolonged, owing to the slow diffusion of the antisep- 
tic into the aqueous nasal secretions. The oil solution is 
sprayed into the nose and throat from an ordinary oil spray, 
preferably one made entirely of glass, thus avoiding the 
necessity of an inhalation chamber. 







Appli- 










cations 


Time of Test, 


Number of 




Treatment Employed 


of Oil 


from Beginning 


Colonies on 






between 


of Treatment 


Agar Plate 






Tests 






I. 


o-5% aq. chloramine-T 








42,240 




followed by 2 % di-chlor- 


1 


30 min. 


9 




amine-T in oil. 





60 min. 





II. 


1.5% dichloramine-T in 








16,104 




oil. 


1 


1 hour 


175 






1 


3.5 hours 


1 









6 hours 


9 


III. 


o-5% aq. chloramine-T 








6,129 




followed by 1.3% di- 


1 


1 hour 


8,960 




chloramine-T in oil. 


4 


4 hours 


87 









20 hours 


1,980 






1 


22 hours 


59 






1 


24 hours 


3 






1 


26 hours 


1 






1 


27.5 hours 


20 



Experience in the treatment of meningococcus carriers 
with 2 per cent dichloramine-T dissolved in chlorinated 
eucalyptol and paraffin (1:4) has as yet been limited, but 
those cases treated have been promptly freed from the me- 
ningococcus and the method appears likely to be used ex- 
tensively. The above table shows the effect of spraying 
the oil solution of dichloramine-T on the normal nose. In 
such experiments it must be recalled that many of the bac- 



THE DISINFECTION OF CARRIERS 103 

teria are much more resistant to antiseptics than the menin- 
gococcus and that spore forms were probably present. 

In the above experiments, the strongest solution was 2 
per cent and the result was very striking. It should be 
understood, however, that the concentration of the antisep- 
tic in the oil bears no simple relation to the strength ac- 
tually applied to the organisms in the aqueous secretions. 
For example, a sample of the oil containing 2 per cent di- 
chloramine-T, when shaken with an equal volume of clear 
nasal secretion, showed that the filtered aqueous portion 
had a concentration of active chlorine corresponding to about 
0.1 per cent dichloramine-T. The chief advantage of using a 
concentrated oil solution lies in prolonging the period through 
which it can serve as a store of active antiseptic. In the 
normal nose, a 2 per cent solution is not exhausted until 
about two hours have elapsed; but when foul discharges 
are present, this period would be curtailed. 

The experience hitherto gained in the treatment of carriers 
with dichloramine-T indicates that the following procedure 
may be advantageously followed : 

First cleanse the nose with normal salt solution, with or 
without the addition of 0.25 per cent chloramine-T, by spray- 
ing, or irrigation. The same chloramine-T solution should 
be used as a gargle. After this preliminary treatment and 
when the augmentation of nasal, secretion has subsided, 
apply the oil solution of dichloramine-T l (2 per cent) with 
an oil " atomizer," endeavoring to reach all the parts with 
an abundant supply of oil. It is not certain that the oil so 
introduced is ever active for more than two hours, so that 
for intensive treatment it should be renewed at the end of 

1 This solution is best prepared by dissolving 0.2 gram of dichloramine-T 
in 2 cc. chlorinated eucalyptol and when all dissolved, adding 8 cc. chlorinated 
paraffin. The solution should be discarded as soon as signs of a definite crys- 
talline sediment appear. Ordinarily it is stable for about three days. It should 
be carefully protected as far as possible from direct exposure to sunlight, as this 
hastens its decomposition. 



104 A HANDBOOK OF ANTISEPTICS 

that time. In any case, it appears important to repeat 
spraying with the oil so that four treatments a day are 
given at about equal intervals. The first few applications 
of oil sometimes occasion sneezing, but the nose appears to 
acquire a tolerance for the treatment, and subsequent applica- 
tions occasion no inconvenience. In cases of acute coryza, 
dichloramine-T applied as described is too irritating to the 
inflamed membrane and its use is not advised. 

In the way of prophylaxis, spraying of the upper air pas- 
sages with a 0.25 per cent solution of chloramine-T in saline, 
or an oil solution of dichloramine-T appears to be followed 
by good results in reducing the incidence of coryza, measles 
and mumps, although it is not possible to cite convincing 
statistical evidence. Colonel Parke's x report on army trans- 
ports covering long voyages in which an inhalatorium of the 
Gordon and Flack type was used, lends support to the view 
that such treatment is useful. The simplicity of these 
methods and the apparent value of the results indicate that 
more extended trials are desirable. 

Among the other antiseptic solutions employed in the 
treatment of meningococcus carriers, reference may be made 
to an iodine and menthol preparation frequently employed 
in England, while Vincent in the French army advocated 
the following mixture as an inhalation: iodine 12 grams, 
potassium iodide 6 grams, guaiacol 2 grams, thymol 0.35 
gram, alcohol 200 grams, used five or six times a day. For 
disinfection of the pharynx, swabbing with a 3 to 5 per cent 
solution of glycerine and iodine is advocated. This would 
appear a decidedly drastic treatment. Sophian has recom- 
mended hydrogen peroxide 1 per cent and argyrol 9 per 
cent, used as a spray, as of value for freeing carriers of me- 
ningococci, although the mixture does not seem to have been 
used extensively in military centres. 

The antiseptic treatment of carriers has been much more 

1 Brit. Med. Journ., Feb. 24, 1917. 



THE DISINFECTION OF WATER 105 

successful in meningococcus cases than with most other infec- 
tions. The reasons for this are doubtless due in large meas- 
ure to the fact that the meningococcus is very readily killed 
by antiseptics and is usually found in situations accessible 
to treatment. The treatment of diphtheria carriers has been 
less successful, though encouraging, while pneumococcus 
carriers give disappointing results. The reasons for this 
are obvious when the pathological conditions are considered. 

II. The Disinfection of Water 

The chemical disinfection on a large scale of contaminated 
or suspected water supplies for civilian military or naval 
use is now almost exclusively effected with substances 
belonging to the chlorine group. 1 When working with 
considerable volumes of water, the choice of reagent is prac- 
tically limited to free chlorine supplied from cylinders of 
the liquefied gas, sodium hypochlorite solutions of known 
concentration, or bleaching powder (chloride of lime). The 
action of these substances is essentially similar, and their 
usefulness depends upon the fact that their addition in such 
quantity that about 0.5 to 1 part of active chlorine per 
million parts of water persists for a short period, is followed 
by an almost complete disappearance of organisms of the 
coli group. Spore forms which are of little hygienic impor- 
tance are not much affected. Preliminary filtration of 
turbid waters is desirable, and occasionally in the case of 
extreme turbid waters, clarifying with some precipitant, 
such as alum followed by alkali, is resorted to. 

The choice of reagent will depend largely upon the actual 
conditions of its employment. When large volumes of 
water are to be treated and liquefied chlorine is available, 

1 An old chemical method of purifying water depended on the use of potassium 
permanganate. Except against the cholera spirillum, it is not very effective, 
and the cost of the material, and the disagreeable appearance and taste of the 
treated water, are serious drawbacks. 



106 A HANDBOOK OF ANTISEPTICS 

this may be the most convenient method, and especially 
if a gas gauge such as the Wallace-Tiernan is employed 
by means of which the desired chlorine concentration in a 
known quantity of water may be easily regulated. Sodium 
hypochlorite solution is preferable to bleaching powder in 
that it is more readily distributed through the water, but 
its varying composition and instability make it less satis- 
factory for field use. For the sterilization of moderate 
volumes of water such as those contained in army water 
carts, e.g. no gallons, bleaching powder has given most 
satisfactory results. 

The successful use of chlorine compounds for the disin- 
fection of water depends largely upon adequate mixing 
with the water; the use of an effective concentration of 
disinfectant such as that already indicated ; and lastly the 
avoidance of excess. The taste and smell of unnecessarily 
highly chlorinated water is objectionable to most people. 
Not infrequently a reducing substance such as sodium 
thiosulphate is added to remove the excess of chlorine, but 
with careful chlorination this should not be necessary. The 
exact amount of disinfectant to be added to water can only 
be fixed with regard to individual supplies of constant com- 
position. The reason for this is that different waters con- 
tain such varying amounts of organic matter and some- 
times ferrous salts or sulphides as well, and the chlorine 
used up in oxidizing these substances will not effect steriliza- 
tion. A slight excess only of the disinfectant must therefore 
be employed, and the proper amount is most readily deter- 
mined by adding a little more {e.g. 0.5 to i per million) than 
the minimum quantity necessary for obtaining a positive 
reaction for active chlorine in the water. The test for active 
chlorine is most easily carried out by adding a few drops each 
of 10 per cent potassium iodide solution and starch, paste to 
about 200 cc. of water and noting whether a blue color de- 
velops. Since starch paste is apt to become mouldy, it is 



THE DISINFECTION OF WATER 107 

often convenient to use zinc iodide (7.5 per cent) in place of 
the potassium salt and to add this directly to the starch 
paste together with two per cent of alcohol. This mix- 
ture keeps well. Various simple devices have been used 
for carrying out this test in the field so that accurate dosage 
may be rapidly determined. It should be remembered 
that the sensitiveness of the starch-iodide reaction for active 
chlorine rapidly diminishes with rise in temperature of the 
water. The speed with which the reaction for active chlorine 
disappears from a treated water depends upon many factors, 
such as the degree of surface exposure, carbon dioxide con- 
centration, the presence or absence of much ammonia in 
the water, and the nature and amount of the organic matter. 
Under most circumstances, an initial concentration of 0.5 
part per million of active chlorine will vanish in a very 
few hours. 

It has been observed that if an addition of ammonia is 
made to the hypochlorite to be used for water sterilization, 
the resulting product monochloramine, NH 2 C1, is still 
more active as a disinfectant than the original hypochlorite, 1 
and its action is more prolonged. Use of this fact has been 
made in the sterilization of the water supplies of several 
large communities with excellent results. A concentration 
of 0.6 part per million of active chlorine, used as mono- 
chloramine, appears to give a considerable margin of 
safety. 

Further details concerning the large scale disinfection of 
water should be sought in books dealing primarily with 
questions of hygiene. 

The Disinfection of Small Quantities of Water. — 
Chlorine gas, bleaching powder or sodium hypochlorite, which 
are so effective for the disinfection of relatively large volumes 
of water, are not suitable for the individual sterilization of 
small quantities such as the contents of an army water bottle. 

1 Rideal, Journ. Roy. Sanitary Inst., 31, p. 33, 1910. 



108 A HANDBOOK OF ANTISEPTICS 

The problem of sterilizing small individual quantities of 
water such as are needed by cavalry or rapidly moving troops 
is a difficult one, for the use of the chlorine antiseptics just 
mentioned is generally impracticable owing to the instability 
of small tablets containing the required minute quantity 
of active disinfectant. In their place acid sulphates of 
the alkali metals have been used to a considerable extent, 
and these will be considered later ; but the superior potency 
of the chlorine antiseptics induced us to try and find suitably 
stable compounds, and one of these which will now be de- 
scribed has met with some success as the result of practical 
trials. 

A number of compounds which at first sight seemed likely 
to be useful had to be discarded either because they were 
ineffective in hard waters or because of difficulties concern- 
ing solubility or stability. The most suitable substance 
that we have as yet found is p-sulphondichloramino- 
benzoic acid, — Cl2N0 2 SC 6 H 4 COOH. It is easily pre- 
pared from cheap, readily available materials, and appears 
to be effective and reasonably stable. The presence of the 
COOH group confers a slight but definite degree of solu- 
bility in water, which is increased by dispensing it with 
alkaline salts such as sodium carbonate or borax. For- 
mulae for the tablets are appended together with details 
of the preparation of the substance and an estimate of its 
cost. Since the systematic name of the disinfectant is 
inconveniently long for ordinary use, we have adopted the 
name "halazone." The abbreviated name gives some indi- 
cation of the character of the compound. 

In the following table are given some of the bacteriological 
results. The technique employed was that in general use 
and requires no special description. Five or ten standard 
drops of the treated w r ater were generally used for plating 
on agar for counting the surviving organisms. Suitable 
controls were invariably carried out. 



THE DISINFECTION OF WATER 



109 



Experiments with p-Sulphondichloraminobenzoic Acid in 
Tablet Form. 



No. 


Water Treated 


Concentra- 
tion of Dis- 




H H 
< P 

'- - 


Surviving 
Organisms 


Q 
W 
> 






infectant 


per c.c. 


s^ 








as 




►J 


i 


Tap water B. coli. 


- 





112,525 




2 


Tap water B. coli. 


1 : 400,000 


15 



178,528 


IO 






1 : 500,000 


15 


987 


IO 


3 


Tap water 5% sewage B. coli. 


1-: 500,000 


30 



1,119,000 


IO 






1 : 175,000 


15 


6,000 


9 


4 


Tap water 5% sewage B. coli. 




30 
60 






1,158,500 


9 
9 






1 : 330,000 


15 


120,064 


7 






1 : 330,000 


30 


9,146 


7 


5 


Deep yellow polluted river water B. 


1 : 330,000 


60 





7 




coli. 





— 


33,152 








1 : 250,000 


20 


10,940 


13 






1 : 250,000 


40 


852 




6 


Tap water B. typhosus. 


1 : 250,000 



60 




1,155,400 




7 


Hard water B. typhosus. 


1 : 225,000 



20 



85,400 


' 17 


8 


Tap water 5% sewage B. typhosus. 


1 : 333,333 



20 



66,017 


14 






1 : 333,333 


20 


242 


14 


9 


Tap water B. paratyphosus A. 


1 : 333,333 



40 



112,000 








1 : 225,000 


20 





17 


io 


Tap water 5% sewage B. paraty- 
phosus A. 





. 


29,400 








1 : 333,333 


20 


15 


12 


ii 


Tap water B. paratyphosus B. 


1 : 333,333 



40 



322,500 








1 : 225,000 


20 


10 


17 






1 : 225,000 


40 








110 i 



A HANDBOOK OF ANTISEPTICS 



Experiments with p-Sulphondichloraminobenzoic Acid in 
Tablet Form. — Continued. 



No. 


Water Treated 


Concentra- 
tion or Dis- 


< 


Surviving 
Organisms 


Q 

w 
> 

ft 






infectant 


PER C.C. 














pa 5 


12 


Tap water 5% sewage B. paraty- 
phosus B. 







130,776 








1 : 333,333 


20 


290 


12 


13 


Tap water V. cholerae. 


1 : 333,333 



40 




13,706 




14 


Tap water 5% Sewage V. Cholerae. 


1 : 450,000 



20 



11,170 


I3J 

13 


15 


Tap water B. dysenteriae (Flexner). 


1-333,333 



20 




66,998 








1 : 450,000 


15 


25 


13 


16 


Tap water 5% sewage B . dysenteriae 
(Flexner) . 


1 : 450,000 



30 




164,864 








1 : 333,333 


20 


38 


13 


17 


Tap water B. dysenteriae (Shiga). 


1 : 333,333 



40 



31,200 








1 : 333,333 


20 


10,934 


II 


18 


Tap water 5% sewage B. dysenteriae 
(Shiga). 


1 : 333,333 



40 



2,108 








1 : 333,333 


20 





13 



From the results in the table it appears that a concentra- 
tion of 1 : 300,000 is sufficient to sterilize an ordinarily heavily 
contaminated water in about thirty minutes. Such a con- 
centration could be relied upon to remove coli, typhoid, 
or cholera organisms. Special experiments showed that the 
substance in tablet form was efficacious when acting on water 
contained in aluminium bottles, although a very trifling 
action on the metal may be observed if tablets are allowed to 
remain undisturbed in long contact with the metal. We 



THE DISINFECTION OF WATER 111 

believe such action to be of no practical moment. The 
concentration of disinfectant given above is just perceptible 
to the taste, especially in warm water containing little or- 
ganic matter, but the water is perfectly palatable. Tea 
brewed with or without the addition of halazone to the 
water cannot be distinguished. One point of advantage 
possessed by the present disinfectant over most hypo- 
chlorite preparations is the fact that the active chlorine is 
less rapidly used up, so that the process of disinfection con- 
tinues for a longer period. 1 

Even in large doses the substance is not toxic (250 milli- 
grams per kilo) and is excreted by rabbits in the urine as 
p-.sulphonamidobenzoic acid. 

Preparation and Properties of Halazone. — The 
starting point in the preparation of the substance is p- 
. toluenesulphonamide, a product which is readily obtained 
by the action of ammonia on p-toluenesulphonic chloride. 
The latter substance is a cheap waste product in the manu- 
facture of saccharine, and is available in relatively large 
quantities. It is also used for the manufacture of chloramine- 
T, and dichloramine-T. 

Toluenesulphonamide is oxidized to p-sulphonamido- 
benzoic acid, and the latter substance on treatment with 
chlorine under suitable conditions gives the desired dichlor- 
amino acid. The reactions may be expressed as follows : 



CH 3 COOH COOH 




S0 2 NH 2 S0 2 NH 2 S0 2 NC1 2 



The experimental details are as follows: Add 250 grams 
commercial sodium dichromate to a mixture of 200 cc. 

1 It may be noted that " halazone" is unsuited for use as a preservative for 
milk or other foodstuffs, or as a general antiseptic. 



112 A HANDBOOK OF ANTISEPTICS 

sulphuric acid and 600 cc. water contained in a 2 liter round 
flask. Then add 100 grams crude toluene-p-sulphonamide 
and heat on a sand bath with reflux condenser for one hour, 
using a small flame *at first as the reaction is vigorous. On 
cooling wash the separated crystals well with cold water, 
and then dissolve them in hot dilute sodium hydroxide in 
slight excess. Filter hot, and add excess of hydrochloric 
acid and when cold filter off the precipitated sulphonamido- 
benzoic acid, wash well with water and dry. The yield is 
about 80 per cent of theory. 

Twenty grams of p-sulphonamidobenzoic acid are dis- 
solved in 200 cc. approximately normal sodium hydroxide 
(2 mols.) warming if necessary. About 200 grams of 
crushed ice is then added, and the mixture saturated with 
a rapid current of chlorine. The reaction is most conven- 
iently carried out in a fairly wide-mouthed flask, which may 
be shaken while the gas is being introduced. If the tempera- 
ture should rise more ice can be added. A white, rather chalky 
precipitate of the dichloramino acid is at once precipitated. 
The acid is filtered off, using suction, well washed with cold 
water, and dried in vacuo on a porous plate. The dry sub- 
stance is practically pure, and may be powdered and preserved 
apparently indefinitely. Prepared by this method, the yield 
of the dichloramino acid is practically the theoretical amount. 

The substance is sparingly soluble in water and in chloro- 
form and insoluble in petroleum. It readily dissolves in 
glacial acetic acid, crystallizing in stout prisms, which melt 
at 213 C. The substance explodes feebly when rapidly 
heated on platinum foil, but, compared with most members 
of the group, is remarkably stable. 

The purity of the compound may be checked by titration 
as follows : 0.1 gram is weighed out, dissolved in 50 per cent 
acetic acid and potassium iodide added. The liberated 
iodine is titrated with decinormal sodium thiosulphate, 
of which 14.8 to 14.9 cc. will be required (see p. 40). 



THE DISINFECTION OF WATER 113 

The dichloramino acid dissolves, apparently without 
change, in excess of cold sodium hydrate solution, and may 
be reprecipitated on addition of acids. With smaller quan- 
tities of sodium hydroxide or with feebly alkaline salts, such 
as phosphates or borates, hydrolysis occurs, with liberation 
of disagreeably smelling compounds of nitrogen and chlorine. 

Dry sodium carbonate or dry borax appear the most satis- 
factory alkaline salts for making tablets containing halazone. 
A convenient formula for tablets weighing ioo to 105 mg. 
is to use sulphondichloraminobenzoic acid 4 per cent, sodium 
carbonate 4 per cent (or dried borax 8 per cent) sodium 
chloride (pure) 92 per cent. The acid should be ground 
up with the dry salt and the sodium carbonate or borax 
added subsequently. The mixture, which must be kept 
perfectly dry, may be passed through a 40 mesh sieve. No 
lubricant or other addition is necessary and should be avoided. 
The strength of the tablets should be tested by dissolving 
in 50 per cent acetic acid and potassium iodide solution, 
and titrating with decinormal sodium thiosulphate as al- 
ready described (p. 40) (1 cc. of N/10 thiosulphate = 0.00675 
gram of the dichloramino acid). They must not be allowed 
to dissolve in water and subsequently titrated, as then de- 
composition occurs. The tablets should be stored in small 
amber glass bottles, to protect them from the action of light. 
Tablets so prepared of the weight mentioned contain about 
4 mg. of the disinfectant, and are suitable for the steriliza- 
tion of a liter or quart of reasonably heavily contaminated 
water. In the case of extreme contamination a second 
tablet may be necessary. 1 

The practical success of the disinfectant depends very 
largely on the stability of the tablet. It appears, as judged 
by several months' observation, that the tablets are quite 
stable enough for practical use. They are certainly more 

1 Halazone tablets can be obtained from Boots, Island Street, Nottingham, 
England, or Abbots of Chicago and doubtless other firms. 



114 



A HANDBOOK OF ANTISEPTICS 



permanent than other similar compounds with which we 
are acquainted, and when kept in amber bottles under or- 
dinary conditions, and at temperatures not exceeding 3o°C, 
less than 5 per cent decomposition was observed in 150 days. 

It is rather difficult to give precise estimates of the cost 
of the finished product, but it is safe to say that the tablets 
could be sold at such a price that 100 gallons of water could 
be sterilized at a cost of one penny. 

Sodium Bisulphate and Other Acid Substances. — The 
injurious effect of acids upon the cholera vibrio and typhoid 
bacillus was pointed out long ago by Koch and Kitasato, 
Stutzen, and others. From Kitasato's experiments it ap- 
peared that from about 0.2 to 0.5 per cent of most acids was 
necessary to free water from typhoid bacilli in a few hours, 
but that a notably lower concentration of sulphuric acid 
was effective. The following table has been compiled by 
Colonel Horrocks, illustrating the effect of various acid 
substances upon typhoid bacilli. 





Per Cents of Reagent 


Acids 


Growth 


Growth 

Restrained 


No Growth 


Sulphuric Acid 

Hydrochloric Acid 

Nitric Acid 

Sulphurous Acid 

Phosphoric Acid 

Acetic Acid 

Formic Acid 

Oxalic Acid 

Lactic Acid 

Tartaric Acid 

Citric Acid 

Malic Acid 

Tannic Acid ....... 

Boric Acid 


0.049 

O.I 
O.I 

0.09 

0.15 

0.2 

0.22 

0.23 

0.27 

0.338 

0.338 

0.338 

1-3 

i-5 


0.065 

0.158 

O.I57 

0.2 

0.224 

0.255 

0.278 

0.285 

0.36 

0.384 

0.384 

0.384 

2.0 


0.08 
0.2 
0.2 
0.28 
0.3 
0.3 
o.34 
0.366 
0.4 
0.476 
0.476 
0.476 
. 1 66 
2.7 



THE DISINFECTION OF WATER 115 

In these experiments broth and gelatine were carefully 
neutralized before adding the substances, then inoculated 
with B. typhosus and kept at room temperature, subcul- 
tures being taken at 4-5 and 10-15 hours. Considerably 
greater disinfecting action was obtained with waters contain- 
ing a minimum of organic matter. 

The possibility of using an acid mixture for the sterilization 
of small quantities of water was put into practical form by the 
suggestion that solid tablets of acid sodium sulphate might 
be employed. From the experiments of Rideal x and Parkes 
it appears that one gram of sodium bisulphate per pint of 
water is adequate for the sterilization of waters moderately 
heavily contaminated with typhoid bacilli in fifteen minutes, 
while very heavily infected waters may require forty-five 
minutes. Under similar conditions B. enteritidis is destroyed 
. about as readily as B. typhosus while the V. cholera is more 
sensitive than either. 

Sodium bisulphate tablets were used by the New Zealand 
contingent in the South African War and subsequently to 
some extent by cavalry and mounted police and other mo- 
bile forces. They are frequently compounded with lemon 
oil and saccharin so that the resulting solution has some 
resemblance to lemonade. The taste of this acid mixture 
is actively disliked by some individuals but well tolerated 
by others. The acid sulphates have the disadvantage of 
attacking metallic water bottles to a marked extent and 
while undoubtedly useful, do not possess the potency or 
security afforded by a disinfectant of the chlorine group. 
The tablets apart from mechanical disintegration have the 
advantage of being indefinitely stable. 

1 Rideal, Disinfection and the Preservation of Food, John Wiley & Sons, 1903, 
P- 376. 



116 A HANDBOOK OF ANTISEPTICS 

III. The Disinfection of Hospital Ships, etc., with 
Sodium Hypochlorite Prepared by Electrolysis of 
Sea Water or Brine 

In the following section an account is given of a relatively 
simple and cheap apparatus for the preparation of consider- 
able quantities of sodium hypochlorite by the electrolysis 
of sea water. The results obtained by the use of hypochlorite 
disinfectant in reducing secondary infections on large hos- 
pital ships carrying many infectious cases, have been suffi- 
ciently striking to justify, perhaps, the inclusion of an 
abridged report on the use of this apparatus. 

Opportunities for the employment of the apparatus are 
by no means restricted to hospital ships for the only requi- 
sites are salt water or brine and a supply of direct current 
electrical energy. 

The preparation of sodium hypochlorite by the electrolysis 
of salt solutions is an old process, and since Watt's initial 
discovery in 1859 innumerable modifications in its mode of 
production have been introduced, chiefly for industrial 
purposes. The deodorizing and germicidal properties of 
the hypochlorites have been known for more than a century, 
and it is impossible to refer to many of the applications 
for hygienic purposes which these substances have found. 

Reference may be made, however, to the well-known 
Hermite process for the sterilization of sewage by electrolytic 
hypochlorite, introduced about 1893 ; to the wide employ- 
ment of hypochlorites for the purification of suspected 
potable waters, and to the many uses as disinfectant 
which hypochlorites have found in sanitary work. Elec- 
trolytic hypochlorite for general disinfection purposes has 
been successfully employed by a great many sanitary 
authorities. 

The apparatus to be described was originally employed 
on the Hospital Ship "Aquitania" and since then has been 



ELECTROLYTIC HYPOCHLORITE 117 

adopted on a number of other ships and coast stations. 1 It 
was first of all necessary to devise an apparatus suitable 
for use on shipboard, then to determine favorable conditions 
for carrying out the electrolysis and to decide on suitable 
methods for applying the product. Nothing essentially 
new is embodied in the design or use of the apparatus but 
many existing electrolyzers are either unsuitable or subject 
to patent restrictions which limit their usefulness. 

Description of Apparatus. — The necessary apparatus 
comprises an electrolytic cell, a reversing switch capable 
of carrying a hundred amperes and some ordinary insulated 
electric cable. A scale drawing of the electrolytic cell is 
given on the following pages* 

The electrolyzer consists of a rectangular box made of 
teak or cedar securely bolted together. It may be coated 
internally with marine glue to protect the wood and to reduce 
the risk of leaking. The interior of the box is divided into 
twenty or preferably twenty-five cells by means of carbon 
plates placed parallel to one another. The plates situated 
at both ends of the box project above the level of the other 
plates to receive the terminals for the introduction of the 
current. The terminal electrodes for convenience are made 
in four pieces placed side by side, while the intermediate 
carbons are made in three parts superimposed vertically 
one upon the other. A copper plate attached to the four 
binding screws at each end insures an even distribution of 
current. 

The carbon plates are separated from each other by strips 
of vulcanite or wood, or glass tubes, and they are kept in 
position by means of a " making up block" and wedges 
placed at one end of the tank. A grooved channel is cut 
along one side of the floor of the wooden tank underneath 
the carbon plates in order to facilitate emptying by means 
of a wooden or vulcanite plug tap inserted in a hole con- 
1 Dakin and Carlisle; Jour. Roy. Army Medical Corps, Feb., 1916. 



118 A HANDBOOK OF ANTISEPTICS 

nected with the floor of the tank. A word must be added 
as to the necessity of employing suitable carbon plates. 
These should be of Acheson graphite prepared by treating 
amorphous carbon in the electric furnace. Ordinary carbon 
plates rapidly deteriorate and cannot be used satisfactorily. 
The tank should be mounted on a rubber mat or on glass 
or porcelain insulators, and securely fixed to a low table so 
that its contents can be run conveniently into a tub placed 
beneath it. 

The necessary electrical connections are made as follows : 
— two wires leading the ship's current (one hundred to one 
hundred and ten volts, direct current) are connected re- 
spectively with the middle pair of binding screws in the 
reversing switch. The two binding screws on both sides 
of the switch are each provided with leading wires which 
are attached to the electrolyzer in such a fashion that the 
two wires leading to either side of the switch are attached 
to opposite ends of the electrolyzer. Alternating current 
cannot be used for the electrolysis but must be transformed. 
Most ships furnish direct current of suitable voltage, namely, 
one hundred to one hundred and ten volts. In the case of the 
voltage being two hundred to two hundred and twenty 
volts, two electrolyzers can be placed in series. The appara- 
tus is extremely simple and there is nothing to get out of 
order. It can be easily operated by an untrained person 
after receiving a few minutes' instruction. The wear and 
tear of the apparatus is extremely small. The carbon plates 
are capable of giving many months' service before renewal 
is necessary. 

The apparatus as described above was made by Messrs. 
Mather and Piatt, Park Works, Manchester. We are in- 
debted to Dr. Edward Hopkinson, a director of the company, 
for much valuable assistance in constructing a practical 
and efficient apparatus. The makers' number for the first 
electrolyzer of this type is G.M. 718/15, and may be usefully 



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ELECTROLYTIC HYPOCHLORITE 119 

quoted for identification purposes in any correspondence 
relative to the purchase of similar apparatus. 

The Operation of Electrolysis. — In order to carry 
out an electrolysis the tank is filled with cold salt water to 
a level approximately one and a half to two inches below 
the upper surface of the carbon plates. The object is to 
avoid excessive current leakage between the cells such as 
would occur if the sea water reached a level above the plates. 
If the ship is rolling, less sea water should be used at each 
operation. A tank of the dimensions given will hold about 
thirty to thirty-five liters, or six to seven gallons. The salt 
water should be cold in order to limit the conversion of 
nypochlorite into useless chlorate consequent upon excessive 
heating. Special experiments made with sea water at 
temperatures varying from 5 C. to 25 C. showed that the 
cell can electrolyze sea water efficiently at any temperature 
likely to be encountered in practice. During the electrolysis 
the temperature of the solution in the cells rises about 1.5 C. 
per minute when a current of about 50 to 70 amperes is 
passing. 

The tank being filled with cold salt water the current is 
turned on by means of the switch. 1 A brisk evolution of 
hydrogen with traces of chlorine is at once noticeable, and 
neutral sodium hypochlorite is formed in the solution. The 
following table gives details of a single typical experiment 
illustrating the course of electrolysis after varying lengths 
of time. It will be noticed that after a few minutes the 
quantity of available chlorine formed per minute steadily 
diminishes. The efficiency of the cell being greatest in the 
early stages of the electrolysis, it is most economical to 
electrolyze for a short time rather than to aim at attaining 
a very high concentration of hypochlorite. Using ordinary 

1 Untrained attendants must be warned against turning the current on before 
filling the tank and against adding salt water to the tank while electrolysis is 
in progress. 



120 



A HANDBOOK OF ANTISEPTICS 



salt water, it is inadvisable to continue the electrolysis for 
more than ten minutes. With a current of 60 to 75 amperes 
at no volts, ten minutes' electrolysis will give a solution 
usually containing a little more than four parts per thousand 
of sodium hypochlorite or of available chlorine. For most 
purposes it is distinctly advantageous to limit the electrolysis 
to five minutes. Under these conditions the efficiency of 
the cell is high and the rise of the temperature of the solution 
is not great — about 7 to 8°. The solution at the end of 
five minutes will contain from 2.0 to 2.7 parts per thousand 
of sodium hypochlorite or available chlorine. 





Available Chlorine 






Time of 










Electrolysis 




Yield per 


Total /Avail- 


Additional Data 




Minutes 


Grams 


Minute Stage 


able Chlorine 








per Liter 


Grams per 
Liter 


Produced 
Grams 






1 


0.41 


0.41 








2 


0.92 


0.51 








3 


1.49 


o.57 




35 liters sea water. 




4 


2.08 


. 0.59 




Temperature at start, 15 


C. 


5 


2.66 


0.58 


141.4 


Temperature at finish, 2 8° 


C. 


6 


3 -09 


0-43 




Temperature rise, 13 C. 




7 


3-37 


0.28 




Mean voltage no i 1 




8 


3-04 


0.27 




Mean amperage, 72. 




9 


3-Qo 


0.26 








10 


4.04 


0.14 









The direction of the current should be reversed from time 
to time by turning the handle of the reversing switch in the 
opposite direction to that previously used. A daily reversal 
is probably sufficient, but it may be done more often with 
advantage. Current reversal keeps the carbon plates free 
from deposits of magnesium hydroxide, which otherwise 
would lower the efficiency of the cell. 



ELECTROLYTIC HYPOCHLORITE 121 

When less current than sixty amperes is taken by the cell, 
the yield of hypochlorite is naturally lower. 

With approximately constant current the yield of hypo- 
chlorite- is remarkably uniform so far as practical purposes 
are concerned. Where higher concentrations of hypochlorite 
are required, they may be obtained by electrolyzing cooled 
brine containing ten or fifteen per cent of salt. 

Apart from a few particles of suspended carbon from the 
electrode, the electrolyzed solution will be found clear, 
bright, and free from sediment. It should be run off into 
wooden, tubs or other receptacles, such as slate or lead-lined 
vats. 

It is desirable to make a few tests of the output of hypo- 
chlorite on installing a new apparatus on board ship, es- 
pecially when the current available differs greatly from that 
indicated. This, however, is a very easy matter. (See 
page 41.) 

Cost of Apparatus and of Electrolytic Hypochlorite. 
— The cost of disinfection with electrolytic chlorine may 
be resolved into two items : (1) the initial cost of the cell, 
and (2) the cost of the current used for electrolyzing the sea 
water. 

From an examination of the results recorded in Table I, 
it is calculated that when electrolyzing 35 liters of sea water 
at a time for five-minute periods, using no volts and 75 
amperes, the power required to produce a kilogram of 
chlorine is approximately eight kilowatt hours. The cost 
of the electrical energy naturally varies enormously accord- 
ing to the conditions of its production — the estimates vary- 
ing from 0.25 pence to 1.5 pence per unit. Taking 0.75 
pence per unit as a fair average, the cost of producing icco 
liters of hypochlorite solution at 1 : 1000 chlorine concen- 
tration works out at sixpence. This is equivalent to three- 
pence per 100 gallons, a figure which for practical purposes 
may be regarded as almost negligible. 



122 A HANDBOOK OF ANTISEPTICS 

The original cost of the cell installed on shipboard is about 
£ 50, and its depreciation is undoubtedly small. The carbon 
plates will last for over a year with constant use and hence 
much longer when intermittently worked, as on board ship. 
Moreover, they can be readily replaced at moderate cost. 
The tank itself, being of teak or cedar wood, can be easily 
repaired in case of need by the ship's carpenters. 

Judging by certain estimates supplied to us, it is calculated 
that on a ship the size of H.M.H.S. " Aquitania " the economy 
in largely replacing expensive coal tar disinfectants, such as 
carbolic acid, cresol, etc., by electrolytic hypochlorite will 
approximately pay for the cost of the cell in the course of a 
single trip of three weeks. 

The Optimum Concentration of the Electrolytic 
Hypochlorite for Disinfectant Purposes in Wards, etc 
— It is a somewhat difficult matter to select the most ad- 
vantageous concentration for the disinfecting fluid. As is 
well known, the hypochlorites possess extraordinarily high 
germicidal action when acting upon organisms suspended 
in pure water. One to two parts per million of hypochlorite 
will effectively kill, in two hours, a moderately large quantity 
of organisms other than spore forms. But this activity is 
much reduced when the hypochlorite acts in the presence of 
much extraneous matter. It is obvious, therefore, that the 
nature and quantity of the organic matter present on the 
soiled surfaces which are to be disinfected is a matter of 
importance. It would appear that a concentration of 
hypochlorite should be chosen, such as will insure an excess 
of hypochlorite on the treated area, for some short time 
after its application. This ideal concentration will of course 
vary with the nature of the surface treated and the amount 
of dirt upon it. Hypochlorite disappears rapidly when spread 
in a thin film on a wooden surface, less rapidly on linoleum, 
still less rapidly on rubber surfaces, while on a clean glass sur- 
face the hypochlorite may persist for thirty-six hours or more. 



ELECTROLYTIC HYPOCHLORITE 123 

Practically speaking, it will be found that a strength of 
about i per iooo sodium hypochlorite or available chlorine, 
when properly applied, will suffice for all ordinary purposes. 
On slightly contaminated smooth surfaces such as glass, 
rubber, and certain composition floors, i per 3000 or even 
less would probably be found sufficient, but no disadvantages 
follow the employment of the stronger solution. Electroly- 
tic hypochlorite at a concentration of 1 per 1000 available 
chlorine can be used freely in the wards for swabbing and 
mopping the floors, walls, latrines, etc., without objectionable 
after-effect. This solution was conveniently obtained by 
taking the electrclyzed sea water through which the current 
had been passed for five minutes, previously described, 
and diluting each bucketful of this liquor with a bucketful 
of fresh salt water. The diluted mixture was kept in a large 
tub from which it was distributed to the wards. 

The germicidal action of this mixture tested against ty- 
phoid organisms under the conditions of the Walker-Rideal 
test, may be calculated from the experiments of Klein, 
Sommerville and Walker, Rideal, and others. When diluted 
20 times {i.e. a chlorine concentration of 1 : 20,000) it is as 
active as 1 per cent phenol. 

Comparative experiments upon the bacteriological con-' 
ditions of floors before treatment with salt water and after 
treatment with hypochlorite at 1 per 1000, showed a huge 
reduction in the number of organisms to follow the appli- 
cations of hypochlorite, while a large reduction followed 
the use of plain sea water. 

The Uses of Electrolytic Hypochlorite as a Dis- 
infectant in Wards, etc. — The hypochlorite solution 
prepared as described in the previous section by mixing 
equal volumes of salt water electrolyzed for five minutes 
with plain salt water, was kept in large wooden tubs at con- 
venient places near the wards on the different decks. Each 
morning after the floors had been brushed, a company of 



124 A HANDBOOK OF ANTISEPTICS 

sanitary orderlies, carrying the solution in wooden buckets, 
mopped every portion of the floor surface of the ward, under 
the beds, up the dividing walls, and in the lavatories. 1 On 
the wooden decks now enclosed and used for wards, the solu- 
tion dries very quickly, while on linoleum, owing to its non- 
porous character, the wet surface takes longer to dry. It is 
desirable, therefore, that on linoleum a too large excess of 
liquid should not be left, or else a slippery moist surface 
persists for an inconveniently long time. It is important 
that the orderlies be instructed to change the disinfectant 
in their buckets at frequent intervals. The redistribution 
of dirt mixed with a little disinfectant of impaired activity 
over the surface of a ward does not constitute cleaning, and 
it is necessary that this is understood by the orderlies. 

In the enteric and dysentery wards hypochlorite disin- 
fectant was placed in the bed-pans before use and its marked 
deodorant action was much appreciated. After cleaning 
the bed-pans with a special spray, they were well rinsed with 
hypochlorite solution. At frequent intervals the disinfectant 
was used for mopping all the surfaces in the latrines, includ- 
ing the door-handles, and from time to time it was poured 
down the waste-pipes, followed by a good flush of salt water. 
When used in this way we have not observed damage greater 
than that caused by other disinfectants, as shown by the 
results of an actual examination of the plumbing in latrines 
where hypochlorite has been used compared with similar 
structures in latrines treated with phenol and other disin- 
fectants. As is well known, intermittent use of electrolyzed 
sea water has been successfully employed on French naval 
ships to render urinal and water-closet traps unobjectionable 
(cf. Gatewood's Naval Hygiene, p. 443, Rebman, 1909). 

1 It may be of interest to mention the fact that the distance of corridors, 
floors, etc., mopped with the disinfectant exceeded three miles in the case of 
the Hospital Ship "Aquitania." In most cases it was possible to make one 
daily application only. 



ELECTROLYTIC HYPOCHLORITE 126 

It has also been tried on British naval ships to a limited 
extent, but we have no details as to the concentration and 
quantity of hypochlorite used, and these are essential points. 

It is worth noting that electrolytic hypochlorite can be 
used in conjunction with soap, and some particularly heavily 
soiled decks were cleaned with this mixture. It is probable, 
however, that reduction in the germicidal action of the 
hypochlorite is followed by the addition of soap. The 
hypochlorite was also used for putting in the spittoons of 
tuberculosis patients and for other similar purposes for which 
disinfectants are commonly employed. 

The results of the free use of hypochlorite were excellent 
and the wards were fresher, cleaner, and freer from objection- 
able odor than they were previous to its use. In the typhoid 
and dysentery wards the results have been particularly 
striking and the absence of odor was most marked. 

The most important effect of all has been the large reduc- 
tion in secondary infection following upon the introduction 
of hypochlorite disinfection on large hospital ships. While 
it would be unfair to refer this fortunate result as exclusively 
due to the hypochlorite, it is generally considered among 
competent observers that the introduction of the hypo- 
chlorite has been an important cause of the improvement. 

Rate of Decomposition or Electrolytic Hypochlorite 
on Keeping. — It is well known that hypochlorite solutions 
prepared by the direct electrolysis of sea water are unstable. 
For practical purposes of disinfection, etc., on shipboard, 
and in other places this is of no moment, since the solution 
is readily prepared as needed and there is no need for lengthy 
storage. On the average the rate of decomposition stored 
in open tubs was about twenty-five per cent of hypochlorite 
present in the twenty-four hours. The rate of decomposi- 
tion is influenced by temperature and by free exposure to 
air. When stored in covered vats the rate of decomposition 
is materially less. Since the electrolytic hypochlorite is so 



126 A HANDBOOK OF ANTISEPTICS 

readily and cheaply prepared it is well to reject solutions 
which have been stored for more than two or three days. 

The Action of Electrolytic Hypochlorite on Va- 
rious Structural Materials. — Comparative experiments . 
showed that clean iron and steel are attacked to a marked 
extent by i per iooo hypochlorite, and the action is relatively 
rapid. Copper is much more slowly attacked, but the action 
is definite. Brass is still less attacked, while aluminium, 
zinc, nickel, and tin are scarcely attacked at all under the 
conditions of the experiments. Lead is the most resistant 
of all the metals tested. Tin plate, nickel plate, galvanized 
iron, are not appreciably attacked if the plating is intact. 
Organic materials such as wood, linoleum, rubber, composi- 
tion floorings of various kinds, take up a certain amount of 
hypochlorite, as would be expected. Wood is the most 
active in this respect and rubber the least, but in no case 
is significant damage done to the material. Wooden tubs 
which have contained strong hypochlorite solutions for long 
periods develop a soft whitish deposit on the surface, but if 
this is not scraped off further action soon ceases. Painted 
wood and metal, at least so far as the materials tested were 
concerned, proved to be very resistant. None of the floor sur- 
faces in the wards on board ship showed significant damage, 
the only complaint being, as already noted, that occasionally 
the saline hypochlorite dried rather slowly on damp days 
on the linoleum surfaces. But this is due to the non-absor- 
bent character of the linoleum, especially when more or less 
saturated with salts from previous applications, rather than 
to any destructive action. These conditions can be easily 
remedied by occasional washing with fresh water. 

It may be noted that sodium hypochlorite prepared in 
the apparatus described can be used successfully both for 
wound treatment and for the disinfection of drinking water. 
Details as to suitable concentrations for these purposes 
have already been given. 



INDEX 



Acetanilide, 76. 

Acids, 71, 114. 

Acriflavine, 9, 63 et seq., 90, 95, 96. 

Alcohol, 72. 

Ambrine, 47. 

Antiseptics, chlorine group of, 17. 

choice of, 9. 

classification of, 2. 

modes of action, 2. 

modes of application, 13. 

stability of, 2. 
Argyrol, 56, 88. 
Aristol, 46. 

Benzoyl acetyl peroxide, 69. 
Benzoyl hydrogen peroxide, 69. 
Bichloride of mercury, see mercuric 

chloride. 
"B.I. P.," 12, 58. 
Bismuth salts, 12, 50 et seq. 
Bleaching powder, 21, 41, 105. 
Borates, 71. 
Borax, 71. 
Boric acid, 71. 
Borsal, 46. 

Brilliant green, 63, 66, 90, 95. 
Bromacetanilide, 76. 
Bromamines, 18. 
Bromnaphthols, 47. 
Burns, 36, 47, 49. 

Carbolic acid, see phenol. 
Carriers, 99. 
Chinosol, 75. 



Chloramine-T, 7, 14, 19, 20, 28 et seq., 
86, 93, 100. 

estimation of, 42 . 
Chloramines, 3, 18 et seq. 
Chlorazene, see chloramine-T. 
Chloride of lime, see bleaching powder. 
Chlorinated eucalyptol, 34, 37. 

lime, see bleaching powder. 

paraffin, 34, 38. 
Chlorine, 19, 105. 

"active," 17, 41, 106. 

group of antiseptics, 17. 
Classification of antiseptics, 2. 
Cloth, disinfection of, 46. 
Corrosive sublimate, see mercuric 

chloride. 
Creolin, 46. 
Cresols, 45. 
Crystal violet, 63, 66. 

Dakin's solution, 8, 23 et seq. 
Daufresne's formula, 23. 
Dibenzoyl peroxide, 69. 
Dichloramine-T, 12, 19, 20, 33 et seq., 
86, 93, 100. 

estimation of, 42. 
Disinfection, hospital ships, of, 116. 

laws governing, 5. 

media, influence of, 7. 

speed of, 10, 83 et seq. 

temperature, influence of, 6. 

Electrolytic hypochlorite, 116. 
Ether, 72. 



127 



128 



INDEX 



Eucalyptol, chlorinated, 34, 37. 
Eupad, 22. 

Eusol, 8, 15, 22, 82, 85, 86, 93. See 
hypochlorous acid. 

Flavine, see acriflavine. 
Formaldehyde, 73. 
Formaline, 73. 

Gauze, antiseptic, 31, 57. 

Halazone, 108 et seq. 

Hexalthyl violet, 63 . 

Hexamethylenetetramine, 73 . 

Hexamethyl violet, 62. 

Hydrogen peroxide, 2, 7, 68, 88. 

Hypobromites, 18. 

Hypochlorites, 3, 12, 19 et seq., 84, see 

sodium hypochlorite. 
Hypochlorous acid, 3, 19 et seq., see 

eusol. 
estimation of, 41. 
Hypoiodites, 18. 

Intravenous injection, 15. 
Iodine, 7, 8, 69, 82, 86. 
Iodoform, 12, 74. 

Leucocytes, activity of, 10, 83. 
Lysol, 45, 82. 

Malachite green, 61, 66, 82, 89, 95. 
Media, influence of, on disinfection, 7, 

80 et seq. 
Mercuric chloride, 4, 7, 52 et seq., 82, 

87. 
Mercury biniodide, 8, 11, 56, 82. 
salts, 50 et seq. 

Naphthol, 47. 
Nasal antisepsis, 99. 

Ozone, 69. 



Paraffin oil, chlorinated, 34, 38. 
Pastes, antiseptic, 15, 32, 45, 50, 58 et 

seq. 
Perborates, 71. 
Permanganates, 74, 105. 
Peroxides, 68, 69. 
Persulphates, 71. 

Phenol, 7, 8, n, 29, 43 et seq., 82, 88. 
Phenol coefficient, 79. 
Phenols, 4, 43 et seq. 
Picric acid, 48. 
Proflavine, 67, 90, 96. 
Proteins, chlorination of, 18. 
Pus, sterilization of, 8. 
Pyxol, 46. 

Quinine, 74. 

Resorcinol, 48. 

Salicylic acid, 7, 46. 
Ships, disinfection of, 116. 
Silver, colloidal, 56. 

fluoride, 51. 

nitrate, 4, 7, 16, 58, 87. 

salts, 50 et seq. 

sodium cyanide, 51. 
Sodium bisulphate, 114. 

hypochlorite, 7, 8, 12, 19 et seq., 82, 
85, 93, 105, 116. 

estimation of, 41. 

thiosulphate, 40. 

toluene-sulphonchloramide, see . 
chloramine-T. 
Sulphonamidobenzoic acid, in. 
Sulphondichloraminobenzoic acid, 108. 

Tablets, for water, 108, 115. 
Temperature, influence on disinfection, 

6. 
Thymol, 46. 
Toluene-sodium-sulphonchloramide, 

see chloramine-T. 



INDEX 



129 



Toluene sulphondichloramine, 
dicMoramine-T. 

Tribromnaphthol, 48. 

Tricresol, 45. 

Topaflavine, see acrirlavine. 



Water, disinfection of, 105. 

Zinc chloride, 57, 58, 88. 
salts, 50 et seq. 
sulphate, 100. 



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Technique of Operations on the Bones, 
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By ROBERT SOUTTER 

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